Methamphetamine is a highly addictive psychostimulant that causes profound damage to the brain and other body organs. Post mortem studies of human tissues have linked the use of this drug to diseases associated with aging, such as coronary atherosclerosis and pulmonary fibrosis, but the molecular mechanism underlying these findings remains unknown. Here we used functional lipidomics and transcriptomics experiments to study abnormalities in lipid metabolism in select regions of the brain and, to a greater extent, peripheral organs and tissues of rats that self-administered methamphetamine. Experiments in various cellular models (primary mouse fibroblasts and myotubes) allowed us to investigate the molecular mechanisms of systemic inflammation and cellular aging related to methamphetamine abuse. We report now that methamphetamine acceleratescellularsenescence and activates transcription of genes involved in cell-cycle control and inflammation by stimulating production of the sphingolipid messenger ceramide. This pathogenic cascade is triggered by reactive oxygen species, likely generated through methamphetamine metabolism via cytochrome P450, and involves the recruitment of nuclear factor-κB (NF-κB) to induce expression of enzymes in the de novo pathway of ceramide biosynthesis. Inhibitors of NF-κB signaling and ceramide formation prevent methamphetamine-induced senescence and systemic inflammation in rats self-administering the drug, attenuating their health deterioration. The results suggest new therapeutic strategies to reduce the adverse consequences of methamphetamine abuse and improve effectiveness of abstinence treatments. PMID:25671639

Methamphetamine is a highly addictive psychostimulant that causes profound damage to the brain and other body organs. Post mortem studies of human tissues have linked the use of this drug to diseases associated with aging, such as coronary atherosclerosis and pulmonary fibrosis, but the molecular mechanism underlying these findings remains unknown. Here we used functional lipidomics and transcriptomics experiments to study abnormalities in lipid metabolism in select regions of the brain and, to a greater extent, peripheral organs and tissues of rats that self-administered methamphetamine. Experiments in various cellular models (primary mouse fibroblasts and myotubes) allowed us to investigate the molecular mechanisms of systemic inflammation and cellular aging related to methamphetamine abuse. We report now that methamphetamine acceleratescellularsenescence and activates transcription of genes involved in cell-cycle control and inflammation by stimulating production of the sphingolipid messenger ceramide. This pathogenic cascade is triggered by reactive oxygen species, likely generated through methamphetamine metabolism via cytochrome P450, and involves the recruitment of nuclear factor-κB (NF-κB) to induce expression of enzymes in the de novo pathway of ceramide biosynthesis. Inhibitors of NF-κB signaling and ceramide formation prevent methamphetamine-induced senescence and systemic inflammation in rats self-administering the drug, attenuating their health deterioration. The results suggest new therapeutic strategies to reduce the adverse consequences of methamphetamine abuse and improve effectiveness of abstinence treatments.

Senescence was originally described from the observation of the limited ability of normal cells to grow in culture, and may be generated by telomere erosion, accumulation of DNA damages, oxidative stress and modulation of oncogenes or tumor suppressor genes. Senescence corresponds to a cellular response aiming to control tumor progression by limiting cell proliferation and thus constitutes an anticancer barrier. Senescence is observed in pre-malignant tumor stages and disappears from malignant tumors. Agents used in standard chemotherapy also have the potential to induce senescence, which may partly explain their therapeutic activities. It is possible to restore senescence in tumors using targeted therapies that triggers telomere dysfunction or reactivates suppressor genes functions, which are essential for the onset of senescence.

Cellularsenescence is an important mechanism for preventing the proliferation of potential cancer cells. Recently, however, it has become apparent that this process entails more than a simple cessation of cell growth. In addition to suppressing tumorigenesis, cellularsenescence might also promote tissue repair and fuel inflammation associated with aging and cancer progression. Thus, cellularsenescence might participate in four complex biological processes (tumor suppression, tumor promotion, aging, and tissue repair), some of which have apparently opposing effects. The challenge now is to understand the senescence response well enough to harness its benefits while suppressing its drawbacks. PMID:21321098

Standard-of-care chemo- or radio-therapy can induce, besides tumor cell death, also tumor cell senescence. While senescence is considered to be a principal barrier against tumorigenesis, senescent cells can survive in the organism for protracted periods of time and they can promote tumor development. Based on this emerging concept, we hypothesized that elimination of such potentially cancer-promoting senescent cells could offer a therapeutic benefit. To assess this possibility, here we first show that tumor growth of proliferating mouse TC-1 HPV-16-associated cancer cells in syngeneic mice becomes accelerated by co-administration of TC-1 or TRAMP-C2 prostate cancer cells made senescent by pre-treatment with the anti-cancer drug docetaxel, or lethally irradiated. Phenotypic analyses of tumor-explanted cells indicated that the observed acceleration of tumor growth was attributable to a protumorigenic environment created by the co-injected senescent and proliferating cancer cells rather than to escape of the docetaxel-treated cells from senescence. Notably, accelerated tumor growth was effectively inhibited by cell immunotherapy using irradiated TC-1 cells engineered to produce interleukin IL-12. Collectively, our data document that immunotherapy, such as the IL-12 treatment, can provide an effective strategy for elimination of the detrimental effects caused by bystander senescent tumor cells in vivo. PMID:27448982

Cellularsenescence happens in 2 steps: cell cycle arrest followed, or sometimes preceded, by gerogenic conversion (geroconversion). Geroconvesrion is a form of growth, a futile growth during cell cycle arrest. It converts reversible arrest to irreversible senescence. Geroconversion is driven by growth-promoting, mitogen-/nutrient-sensing pathways such as mTOR. Geroconversion leads to hyper-secretory, hypertrophic and pro-inflammatory cellular phenotypes, hyperfunctions and malfunctions. On organismal level, geroconversion leads to age-related diseases and death. Rapamycin, a gerosuppressant, extends life span in diverse species from yeast to mammals. Stress–and oncogene-induced acceleratedsenescence, replicative senescence in vitro and life-long cellular aging in vivo all can be described by 2-step model. PMID:25483060

The cellularsenescence definition comes to the fact of cells irreversible proliferation disability. Besides the cell cycle arrest, senescent cells go through some morphological, biochemical, and functional changes which are the signs of cellularsenescence. The senescent cells (including replicative senescence and stress-induced premature senescence) of all the tissues look alike. They are metabolically active and possess the set of characteristics in vitro and in vivo, which are known as biomarkers of aging and cellularsenescence. Among biomarkers of cellularsenescence telomere shortening is a rather elegant frequently used biomarker. Validity of telomere shortening as a marker for cellularsenescence is based on theoretical and experimental data. PMID:26805432

The cellularsenescence definition comes to the fact of cells irreversible proliferation disability. Besides the cell cycle arrest, senescent cells go through some morphological, biochemical, and functional changes which are the signs of cellularsenescence. The senescent cells (including replicative senescence and stress-induced premature senescence) of all the tissues look alike. They are metabolically active and possess the set of characteristics in vitro and in vivo, which are known as biomarkers of aging and cellularsenescence. Among biomarkers of cellularsenescence telomere shortening is a rather elegant frequently used biomarker. Validity of telomere shortening as a marker for cellularsenescence is based on theoretical and experimental data.

Autophagy and the ubiquitin–proteasome pathway (UPP) are the major protein degradation systems in eukaryotic cells. Whereas the former mediate a bulk nonspecific degradation, the UPP allows a rapid degradation of specific proteins. Both systems have been shown to play a role in tumorigenesis, and the interest in developing therapeutic agents inhibiting protein degradation is steadily growing. However, emerging data point to a critical role for autophagy in cellularsenescence, an established tumor suppressor mechanism. Recently, a selective protein degradation process mediated by the UPP was also shown to contribute to the senescence phenotype. This process is tightly regulated by E3 ubiquitin ligases, deubiquitinases, and several post-translational modifications of target proteins. Illustrating the complexity of UPP, more than 600 human genes have been shown to encode E3 ubiquitin ligases, a number which exceeds that of the protein kinases. Nevertheless, our knowledge of proteasome-dependent protein degradation as a regulated process in cellular contexts such as cancer and senescence remains very limited. Here we discuss the implications of protein degradation in senescence and attempt to relate this function to the protein degradation pattern observed in cancer cells. PMID:24866342

Cellularsenescence has been proposed to play critical roles in tumor suppression and organismal aging, but the molecular mechanism of senescence remains incompletely understood. Here we report that a putative lysosomal carbohydrate efflux transporter, Spinster, induces cellularsenescence in human primary fibroblasts. Administration of d-galactose synergistically enhanced Spinster-induced senescence and this synergism required the transporter activity of Spinster. Intracellular d-galactose is metabolized to galactose-1-phosphate by galactokinase. Galactokinase-deficient fibroblasts, which accumulate intracellular d-galactose, displayed increased baseline senescence. Senescence of galactokinase-deficient fibroblasts was further enhanced by d-galactose administration and was diminished by restoration of wild-type galactokinase expression. Silencing galactokinase in normal fibroblasts also induced senescence. These results suggest a role for intracellular galactose in the induction of cellularsenescence.

Cellular replicative senescence is a major contributing factor to aging and to the development and progression of aging-associated diseases. In this study, we sought to determine viral replication efficiency of influenza virus (IFV) and Varicella Zoster Virus (VZV) infection in senescent cells. Primary human bronchial epithelial cells (HBE) or human dermal fibroblasts (HDF) were allowed to undergo numbers of passages to induce replicative senescence. Induction of replicative senescence in cells was validated by positive senescence-associated β-galactosidase staining. Increased susceptibility to both IFV and VZV infection was observed in senescent HBE and HDF cells, respectively, resulting in higher numbers of plaque formation, along with the upregulation of major viral antigen expression than that in the non-senescent cells. Interestingly, mRNA fold induction level of virus-induced type I interferon (IFN) was attenuated by senescence, whereas IFN-mediated antiviral effect remained robust and potent in virus-infected senescent cells. Additionally, we show that a longevity-promoting gene, sirtuin 1 (SIRT1), has antiviral role against influenza virus infection. In conclusion, our data indicate that enhanced viral replication by cellularsenescence could be due to senescence-mediated reduction of virus-induced type I IFN expression. PMID:27799874

Idiopathic pulmonary fibrosis (IPF) is a fatal disease characterized by interstitial remodelling, leading to compromised lung function. Cellularsenescence markers are detectable within IPF lung tissue and senescent cell deletion rejuvenates pulmonary health in aged mice. Whether and how senescent cells regulate IPF or if their removal may be an efficacious intervention strategy is unknown. Here we demonstrate elevated abundance of senescence biomarkers in IPF lung, with p16 expression increasing with disease severity. We show that the secretome of senescent fibroblasts, which are selectively killed by a senolytic cocktail, dasatinib plus quercetin (DQ), is fibrogenic. Leveraging the bleomycin-injury IPF model, we demonstrate that early-intervention suicide-gene-mediated senescent cell ablation improves pulmonary function and physical health, although lung fibrosis is visibly unaltered. DQ treatment replicates benefits of transgenic clearance. Thus, our findings establish that fibrotic lung disease is mediated, in part, by senescent cells, which can be targeted to improve health and function. PMID:28230051

Idiopathic pulmonary fibrosis (IPF) is a fatal disease characterized by interstitial remodelling, leading to compromised lung function. Cellularsenescence markers are detectable within IPF lung tissue and senescent cell deletion rejuvenates pulmonary health in aged mice. Whether and how senescent cells regulate IPF or if their removal may be an efficacious intervention strategy is unknown. Here we demonstrate elevated abundance of senescence biomarkers in IPF lung, with p16 expression increasing with disease severity. We show that the secretome of senescent fibroblasts, which are selectively killed by a senolytic cocktail, dasatinib plus quercetin (DQ), is fibrogenic. Leveraging the bleomycin-injury IPF model, we demonstrate that early-intervention suicide-gene-mediated senescent cell ablation improves pulmonary function and physical health, although lung fibrosis is visibly unaltered. DQ treatment replicates benefits of transgenic clearance. Thus, our findings establish that fibrotic lung disease is mediated, in part, by senescent cells, which can be targeted to improve health and function.

Cellularsenescence is a potent anti-cancer mechanism that arrests the proliferation of mitotically competent cells to prevent malignant transformation. Senescent cells accumulate with age in a variety of human and mouse tissues where they express a complex ‘senescence-associated secretory phenotype’ (SASP). The SASP includes many pro-inflammatory cytokines, chemokines, growth factors and proteases that have the potential to cause or exacerbate age-related pathology, both degenerative and hyperplastic. While cellularsenescence in peripheral tissues has recently been linked to a number of age-related pathologies, its involvement in brain aging is just beginning to be explored. Recent data generated by several laboratories suggest both aging and age-related neurodegenerative diseases are accompanied by an increase in SASP-expressing senescent cells of non-neuronal origin in the brain. Moreover, this increase correlates with neurodegeneration. Senescent cells in the brain could therefore constitute novel therapeutic targets for treating age-related neuropathologies. PMID:25281806

Cellularsenescence has been widely recognized as a tumor suppressing mechanism that acts as a barrier to cancer development after oncogenic stimuli. A prominent in vivo model of the senescence barrier is represented by nevi, which are composed of melanocytes that, after an initial phase of proliferation induced by activated oncogenes (most commonly BRAF), are blocked in a state of cellularsenescence. Transformation to melanoma occurs when genes involved in controlling senescence are mutated or silenced and cells reacquire the capacity to proliferate. Pirin (PIR) is a highly conserved nuclear protein that likely functions as a transcriptional regulator whose expression levels are altered in different types of tumors. We analyzed the expression pattern of PIR in adult human tissues and found that it is expressed in melanocytes and has a complex pattern of regulation in nevi and melanoma: it is rarely detected in mature nevi, but is expressed at high levels in a subset of melanomas. Loss of function and overexpression experiments in normal and transformed melanocytic cells revealed that PIR is involved in the negative control of cellularsenescence and that its expression is necessary to overcome the senescence barrier. Our results suggest that PIR may have a relevant role in melanoma progression. PMID:21514450

Reactive oxygen species (ROS) were generated in all oxygen-utilizing organisms. Peroxiredoxin II (Prx II) as one of antioxidant enzymes may play a protective role against the oxidative damage caused by ROS. In order to define the role of Prx II in organismal aging, we evaluated cellularsenescence in Prx II(-/-) mouse embryonic fibroblast (MEF). As compared to wild type MEF, cellularsenescence was accelerated in Prx II(-/-) MEF. Senescence-associated (SA)-beta-galactosidase (Gal)-positive cell formation was about 30% higher in Prx II(-/-) MEF. N-Acetyl-l-cysteine (NAC) treatment attenuated SA-beta-Gal-positive cell formation. Prx II(-/-) MEF exhibited the higher G2/M (41%) and lower S (1.6%) phase cells as compared to 24% and 7.3% [corrected] in wild type MEF, respectively. A high increase in the p16 and a slight increase in the p21 and p53 levels were detected in PrxII(-/-) MEF cells. The cellularsenescence of Prx II(-/-) MEF was correlated with the organismal aging of Prx II(-/-) mouse skin. While extracellular signal-regulated kinase (ERK) and p38 activation was detected in Prx II(-/-) MEF, ERK and c-Jun N-terminal kinase (JNK) activation was detected in Prx II(-/-) skin. These results suggest that Prx II may function as an enzymatic antioxidant to prevent cellularsenescence and skin aging.

Proteasome plays fundamental roles in the removal of oxidized proteins and in the normal degradation of short-lived proteins. Previously we have provided evidences that the impairment in proteasome observed during the replicative senescence of human fibroblasts has significant effects on MAPK signaling, proliferation, life span, senescent phenotype and protein oxidative status. These studies have demonstrated that proteasome inhibition and replicative senescence caused accumulation of intracellular protein carbonyl content. In this study, we have investigated the mechanisms by which proteasome dysfunction modulates protein oxidation during cellularsenescence. The results indicate that proteasome inhibition during replicative senescence have significant effects on the intra and extracellular ROS production in vitro. The data also show that ROS impaired the proteasome function, which is partially reversible by antioxidants. Increases in ROS after proteasome inhibition correlated with a significant negative effect on the activity of most mitochondrial electron transporters. We propose that failures in proteasome during cellularsenescence lead to mitochondrial dysfunction, ROS production and oxidative stress. Furthermore, it is likely that changes in proteasome dynamics could generate a pro-oxidative condition at the immediate extracellular microenvironment that could cause tissue injury during aging, in vivo. PMID:17976388

A confounding aspect of biological ageing is the nature and role of senescent cells. It is unclear whether the three major types of cellularsenescence, namely replicative senescence, oncogene-induced senescence and DNA damage-induced senescence are descriptions of the same phenomenon instigated by different sources, or if each of these is distinct, and how they are associated with ageing. Recently, we devised an epigenetic clock with unprecedented accuracy and precision based on very specific DNA methylation changes that occur in function of age. Using primary cells, telomerase-expressing cells and oncogene-expressing cells of the same genetic background, we show that induction of replicative senescence (RS) and oncogene-induced senescence (OIS) are accompanied by ageing of the cell. However, senescence induced by DNA damage is not, even though RS and OIS activate the cellular DNA damage response pathway, highlighting the independence of senescence from cellular ageing. Consistent with this, we observed that telomerase-immortalised cells aged in culture without having been treated with any senescence inducers or DNA-damaging agents, re-affirming the independence of the process of ageing from telomeres and senescence. Collectively, our results reveal that cellular ageing is distinct from cellularsenescence and independent of DNA damage response and telomere length.

Cellularsenescence is considered as an important tumor-suppressive mechanism. Here, we demonstrated that heparan sulfate (HS) prevents cellularsenescence by fine-tuning of the fibroblast growth factor receptor (FGFR) signaling pathway. We found that depletion of 3′-phosphoadenosine 5′-phosphosulfate synthetase 2 (PAPSS2), a synthetic enzyme of the sulfur donor PAPS, led to premature cell senescence in various cancer cells and in a xenograft tumor mouse model. Sodium chlorate, a metabolic inhibitor of HS sulfation also induced a cellularsenescence phenotype. p53 and p21 accumulation was essential for PAPSS2-mediated cellularsenescence. Such senescence phenotypes were closely correlated with cell surface HS levels in both cancer cells and human diploid fibroblasts. The determination of the activation of receptors such as FGFR1, Met, and insulin growth factor 1 receptor β indicated that the augmented FGFR1/AKT signaling was specifically involved in premature senescence in a HS-dependent manner. Thus, blockade of either FGFR1 or AKT prohibited p53 and p21 accumulation and cell fate switched from cellularsenescence to apoptosis. In particular, desulfation at the 2-O position in the HS chain contributed to the premature senescence via the augmented FGFR1 signaling. Taken together, we reveal, for the first time, that the proper status of HS is essential for the prevention of cellularsenescence. These observations allowed us to hypothesize that the FGF/FGFR signaling system could initiate novel tumor defenses through regulating premature senescence. PMID:26250908

Senescent cells accumulate in normal tissues with advancing age and arise by long-term culture of primary cells. Senescence develops following exposure to a range of stress-causing agents and broadly influences the physiology and pathology of tissues, organs, and systems in the body. While many proteins are known to control senescence, numerous noncoding (nc)RNAs are also found to promote or repress the senescent phenotype. Here, we review the regulatory ncRNAs (primarily microRNAs and lncRNAs) identified to-date as key modulators of senescence. We highlight the major senescent pathways (p53/p21 and pRB/p16), as well as the senescence-associated secretory phenotype (SASP) and other senescence-associated events governed by ncRNAs, and discuss the importance of understanding comprehensively the ncRNAs implicated in cell senescence. PMID:26331977

The DNA damage response (DDR) is activated upon DNA damage generation to promote DNA repair and inhibit cell cycle progression in the presence of a lesion. Cellularsenescence is a permanent cell cycle arrest characterized by persistent DDR activation. However, some reports suggest that DDR activation is a feature only of early cellularsenescence that is then lost with time. This challenges the hypothesis that cellularsenescence is caused by persistent DDR activation. To address this issue, we studied DDR activation dynamics in senescent cells. Here we show that normal human fibroblasts retain DDR markers months after replicative senescence establishment. Consistently, human fibroblasts from healthy aged donors display markers of DDR activation even three years in culture after entry into replicative cellularsenescence. However, by extending our analyses to different human cell strains, we also observed an apparent DDR loss with time following entry into cellularsenescence. This though correlates with the inability of these cell strains to survive in culture upon replicative or irradiation-induced cellularsenescence. We propose a model to reconcile these results. Cell strains not suffering the prolonged in vitro culture stress retain robust DDR activation that persists for years, indicating that under physiological conditions persistent DDR is causally involved in senescence establishment and maintenance. However, cell strains unable to maintain cell viability in vitro, due to their inability to cope with prolonged cell culture-associated stress, show an only-apparent reduction in DDR foci which is in fact due to selective loss of the most damaged cells.

Replicative senescence (RS) that limits the proliferating potential of normal eukaryotic cells occurs either by a cell-division counting mechanism linked to telomere erosion or prematurely through induction by cell stressors such as oncogene hyper-activation. However, there is evidence that RS also occurs by a stochastic process that is independent of number of cell divisions or cellular stress and yet it leads to a highly-stable, non-reversible post-mitotic state that may be long-lasting and that such a process is widely represented among higher eukaryotes. Here I present and discuss evidence that the interactions between DNA and the nuclear substructure, commonly known as the nuclear matrix, define a higher-order structure within the cell nucleus that following thermodynamic constraints, stochastically evolves towards maximum stability, thus becoming limiting for mitosis to occur. It is suggested that this process is responsible for ultimate replicative senescence and yet it is compatible with long-term cell survival. PMID:20157542

Vascular endothelial growth factor (VEGF) inhibitors, such as bevacizumab, have improved outcomes in metastatic colorectal cancer (CRC). Recent studies have suggested that VEGF can delay the onset of cellularsenescence in human endothelial cells. As VEGF receptors are known to be upregulated in CRC, we hypothesized that VEGF inhibition may directly influence cellularsenescence in this disease. In our study, we observed that treatment with bevacizumab caused a significant increase (p < 0.05) in cellularsenescence in vitro in several CRC cells, such as MIP101, RKO, SW620 and SW480 cells, compared to untreated or human IgG-treated control cells. Similar results were also obtained from cells treated with a VEGFR2 kinase inhibitor Ki8751. In vivo, cellularsenescence was detected in MIP101 tumor xenografts from 75% of mice treated with bevacizumab, while cellularsenescence was undetectable in xenografts from mice treated with saline or human IgG (p < 0.05). Interestingly, we also observed that the proportion of senescent cells in colon cancer tissues obtained from patients treated with bevacizumab was 4.4-fold higher (p < 0.01) than those of untreated patients. To understand how VEGF inhibitors may regulate cellularsenescence, we noted that among the two important regulators of senescent growth arrest of tumor cells, bevacizumab-associated increase in cellularsenescence coincided with an upregulation of p16 but appeared to be independent of p53. siRNA silencing of p16 gene in MIP101 cells suppressed bevacizumab-induced cellularsenescence, while silencing of p53 had no effect. These findings demonstrate a novel antitumor activity of VEGF inhibitors in CRC, involving p16.

Cellularsenescence is an established tumour-suppressive mechanism that prevents the proliferation of premalignant cells. However, several lines of evidence show that senescent cells, which often persist in vivo, can also promote tumour progression in addition to other age-related pathologies via the senescence-associated secretory phenotype (SASP). Moreover, new insights suggest the SASP can facilitate tissue repair. Here, we review the beneficial and detrimental roles of senescent cells, highlighting conditions under which the senescence response does and does not promote pathology, particularly cancer. By better understanding the context-dependent effects of cellularsenescence, it may be feasible to limit its detrimental properties while preserving its beneficial effects, and develop novel therapeutic strategies to prevent or treat cancer and possibly other age-associated diseases. PMID:27140310

Cells continually experience stress and damage from exogenous and endogenous sources, and their responses range from complete recovery to cell death. Proliferating cells can initiate an additional response by adopting a state of permanent cell-cycle arrest that is termed cellularsenescence. Understanding the causes and consequences of cellularsenescence has provided novel insights into how cells react to stress, especially genotoxic stress, and how this cellular response can affect complex organismal processes such as the development of cancer and ageing.

Cellularsenescence is a fundamental aging mechanism that has been implicated in many age-related diseases and is a significant cause of tissue dysfunction. Accumulation of senescent cells occurs during aging and is also seen in the context of obesity and diabetes. Senescent cells may play a role in type 2 diabetes pathogenesis through direct impact on pancreatic β-cell function, senescence-associated secretory phenotype (SASP)-mediated tissue damage, and involvement in adipose tissue dysfunction. In turn, metabolic and signaling changes seen in diabetes, such as high circulating glucose, altered lipid metabolism, and growth hormone axis perturbations, can promote senescent cell formation. Thus, senescent cells might be part of a pathogenic loop in diabetes, as both a cause and consequence of metabolic changes and tissue damage. Therapeutic targeting of a basic aging mechanism such as cellularsenescence may have a large impact on disease pathogenesis and could be more effective in preventing the progression of diabetes complications than currently available therapies that have limited impact on already existing tissue damage. Therefore, senescent cells and the SASP represent significant opportunities for advancement in the prevention and treatment of type 2 diabetes and its complications.

Alteration of seasonal phenology cues due to climate change has led to changes in the onset and duration of the growing season. While photoperiod often acts as an ultimate control on phenological events, recent studies have shown that environmental cues such as temperature and soil water content can modify the direction and rate of senescence processes. Warmer temperatures have resulted in an observed trend towards delayed senescence across temperate latitudes. However, Arctic regions are characterized by extreme seasonality and rapidly decreasing photoperiod, and consequently senescence may not shift as climate warms. We monitored the timing of Arctic plant community senescence for three years under the framework of an experimental manipulation that altered seasonal phenological cues through warming and earlier snowmelt. Alternative models of senescence were tested to determine if microclimate (air temperature, soil temperature, and soil moisture) or start of season phenology affect the timing and rate of community senescence. We found that all three microclimate predictors contributed to explaining variation in timing of senescence, suggesting that photoperiod is not the sole control on timing of senescence in Arctic plant communities. Rather, increased air and soil temperatures along with drier soil conditions, led to acceleration in the onset of senescence at a community level. Our data suggest that (1) multiple climate drivers predict timing of plant community senescence, and (2) climate change could result in a shorter peak season due to earlier onset of senescence, which would decrease the potential carbon uptake in moist acidic tundra.

Population aging simultaneously highlights the remarkable advances in science, medicine, and public policy, and the formidable challenges facing society. Indeed, aging is the primary risk factor for many of the most common chronic diseases and frailty, which have profound social and economic costs. Population aging also reveals an opportunity; that is, interventions to disrupt the fundamental biology of aging could significantly delay the onset of age-related conditions as a group, and as a result, extend healthy lifespan, or healthspan. There is now considerable evidence that cellularsenescence is an underlying mechanism of aging and age-related conditions. Cellularsenescence is a process in which cells lose the ability to divide and damage neighboring cells by the factors they secrete, collectively referred to as the senescence-associated secretory phenotype (SASP). Herein, we discuss the concept of cellularsenescence, review the evidence that implicates cellularsenescence and the SASP in age-related deterioration, hyperproliferation, and inflammation, and propose that this underlying mechanism of aging may play a fundamental role in the biology of frailty. PMID:26485647

Population aging simultaneously highlights the remarkable advances in science, medicine, and public policy, and the formidable challenges facing society. Indeed, aging is the primary risk factor for many of the most common chronic diseases and frailty, which result in profound social and economic costs. Population aging also reveals an opportunity, i.e. interventions to disrupt the fundamental biology of aging could significantly delay the onset of age-related conditions as a group, and, as a result, extend the healthy life span, or health span. There is now considerable evidence that cellularsenescence is an underlying mechanism of aging and age-related conditions. Cellularsenescence is a process in which cells lose the ability to divide and damage neighboring cells by the factors they secrete, collectively referred to as the senescence-associated secretory phenotype (SASP). Herein, we discuss the concept of cellularsenescence, review the evidence that implicates cellularsenescence and SASP in age-related deterioration, hyperproliferation, and inflammation, and propose that this underlying mechanism of aging may play a fundamental role in the biology of frailty.

Cellularsenescence is thought to play a major role in age-related diseases, which cause nearly 67% of all human deaths worldwide. Recent research in mice showed that exercising mice had higher levels of telomerase, an enzyme that helps maintain telomere length, than nonexercising mice. A commonly used model for biological aging was proposed by Penna. I propose a modification of the Penna model that incorporates cellularsenescence and find an analytical steady-state solution following Coe, Mao, and Cates [Phys. Rev. Lett.PRLTAO0031-900710.1103/PhysRevLett.89.288103 89, 288103 (2002)]. I find that models corresponding to delayed cellularsenescence have younger populations that live longer. I fit the model to the United Kingdom's death distribution, which the original Penna model cannot do.

Epidemiological studies have shown that age is the chief risk factor for lifestyle-related diseases such as cardiovascular disease and diabetes, but the molecular mechanisms that underlie the increase in the risk of such diseases conferred by aging remain unclear. Recently, genetic analyses using various animal models have identified molecules that are crucial for aging. These include components of the DNA repair system, the tumor suppressor pathway, the telomere maintenance system, the insulin/Akt pathway, and other metabolic pathways. Interestingly, most of the molecules that influence the phenotypic changes of aging also regulate cellularsenescence, suggesting a causative link between cellularsenescence and aging. This review examines the hypothesis that cellularsenescence might contribute to lifestyle-related disease.

In this paper we present cellularsenescence as the ultimate driver of the aging process, as a "causal nexus" that bridges microscopic subcellular damage with the phenotypic, macroscopic effect of aging. It is important to understand how the various types of subcellular damage correlated with the aging process lead to the larger, visible effects of anatomical aging. While it has always been assumed that subcellular damage (cause) results in macroscopic aging (effect), the bridging link between the two has been hard to define. Here, we propose that this bridge, which we term the "causal nexus", is in fact cellularsenescence. The subcellular damage itself does not directly cause the visible signs of aging, but rather, as the damage accumulates and reaches a critical mass, cells cease to proliferate and acquire the deleterious "senescence-associated secretory phenotype" (SASP) which then leads to the macroscopic consequences of tissue breakdown to create the physiologically aged phenotype. Thus senescence is a precondition for anatomical aging, and this explains why aging is a gradual process that remains largely invisible during most of its progression. The subcellular damage includes shortening of telomeres, damage to mitochondria, aneuploidy, and DNA double-strand breaks triggered by various genetic, epigenetic, and environmental factors. Damage pathways acting in isolation or in concert converge at the causal nexus of cellularsenescence. In each species some types of damage can be more causative than in others and operate at a variable pace; for example, telomere erosion appears to be a primary cause in human cells, whereas activation of tumor suppressor genes is more causative in rodents. Such species-specific mechanisms indicate that despite different initial causes, most of aging is traced to a single convergent causal nexus: senescence. The exception is in some invertebrate species that escape senescence, and in non-dividing cells such as neurons, where

In this paper we present cellularsenescence as the ultimate driver of the aging process, as a “causal nexus” that bridges microscopic subcellular damage with the phenotypic, macroscopic effect of aging. It is important to understand how the various types of subcellular damage correlated with the aging process lead to the larger, visible effects of anatomical aging. While it has always been assumed that subcellular damage (cause) results in macroscopic aging (effect), the bridging link between the two has been hard to define. Here, we propose that this bridge, which we term the “causal nexus”, is in fact cellularsenescence. The subcellular damage itself does not directly cause the visible signs of aging, but rather, as the damage accumulates and reaches a critical mass, cells cease to proliferate and acquire the deleterious “senescence-associated secretory phenotype” (SASP) which then leads to the macroscopic consequences of tissue breakdown to create the physiologically aged phenotype. Thus senescence is a precondition for anatomical aging, and this explains why aging is a gradual process that remains largely invisible during most of its progression. The subcellular damage includes shortening of telomeres, damage to mitochondria, aneuploidy, and DNA double-strand breaks triggered by various genetic, epigenetic, and environmental factors. Damage pathways acting in isolation or in concert converge at the causal nexus of cellularsenescence. In each species some types of damage can be more causative than in others and operate at a variable pace; for example, telomere erosion appears to be a primary cause in human cells, whereas activation of tumor suppressor genes is more causative in rodents. Such species-specific mechanisms indicate that despite different initial causes, most of aging is traced to a single convergent causal nexus: senescence. The exception is in some invertebrate species that escape senescence, and in non-dividing cells such as neurons

Replicative cellularsenescence is a fundamental biological process characterized by an irreversible arrest of proliferation. Senescent cells accumulate a variety of epigenetic changes, but the three-dimensional (3D) organization of their chromatin is not known. We applied a combination of whole-genome chromosome conformation capture (Hi-C), fluorescence in situ hybridization, and in silico modeling methods to characterize the 3D architecture of interphase chromosomes in proliferating, quiescent, and senescent cells. Although the overall organization of the chromatin into active (A) and repressive (B) compartments and topologically associated domains (TADs) is conserved between the three conditions, a subset of TADs switches between compartments. On a global level, the Hi-C interaction matrices of senescent cells are characterized by a relative loss of long-range and gain of short-range interactions within chromosomes. Direct measurements of distances between genetic loci, chromosome volumes, and chromatin accessibility suggest that the Hi-C interaction changes are caused by a significant reduction of the volumes occupied by individual chromosome arms. In contrast, centromeres oppose this overall compaction trend and increase in volume. The structural model arising from our study provides a unique high-resolution view of the complex chromosomal architecture in senescent cells. PMID:26989773

Idiopathic pulmonary fibrosis (IPF) is an age-related fatal disease with unknown etiology and no effective treatment. In this study, we show that primary cultures of fibroblasts derived from lung biopsies of IPF patients exhibited (i) accelerated replicative cellularsenescence (CS); (ii) high resistance to oxidative-stress-induced cytotoxicity or CS; (iii) a CS-like morphology (even at the proliferative phase); and (iv) rapid accumulation of senescent cells expressing the myofibroblast marker α-SMA. Our findings suggest that CS could serve as a bridge connecting lung aging and its quite frequent outcome -- pulmonary fibrosis, and be an important player in the disease progression. Consequently, targeting senescent cells offers the potential of being a promising therapeutic approach. PMID:26399448

Half a century is fast approaching since Hayflick and colleagues formally described the limited ability of normal human cells to proliferate in culture (Hayflick and Moorhead, 1961). This finding--that normal somatic cells, in contrast to cancer cells, cannot divide indefinitely--challenged the prevailing idea that cells from mortal multicellular organisms were intrinsically ''immortal'' (Carrell, 1912). It also spawned two hypotheses, essential elements of which persist today. The first held that the restricted proliferation of normal cells, now termed cellularsenescence, suppresses cancer (Hayflick, 1965; Sager, 1991; Campisi, 2001). The second hypothesis, as explained in the article by Lorenzini et al., suggested that the limited proliferation of cells in culture recapitulated aspects of organismal aging (Hayflick, 1965; Martin, 1993). How well have these hypotheses weathered the ensuing decades? Before answering this question, we first consider current insights into the causes and consequences of cellularsenescence. Like Lorenzini et al., we limit our discussion to mammals. We also focus on fibroblasts, the cell type studied by Lorenzini et al., but consider other types as well. We suggest that replicative capacity in culture is not a straightforward assessment, and that it correlates poorly with both longevity and body mass. We speculate this is due to the malleable and variable nature of replicative capacity, which renders it an indirect metric of qualitative and quantitative differences among cells to undergo senescence, a response that directly alters cellular phenotype and might indirectly alter tissue structure and function.

Sirtuin 1 (SIRT1) regulates inflammation, aging (life span and health span), calorie restriction/energetics, mitochondrial biogenesis, stress resistance, cellularsenescence, endothelial functions, apoptosis/autophagy, and circadian rhythms through deacetylation of transcription factors and histones. SIRT1 level and activity are decreased in chronic inflammatory conditions and aging, in which oxidative stress occurs. SIRT1 is regulated by a NAD(+)-dependent DNA repair enzyme, poly(ADP-ribose) polymerase-1 (PARP1), and subsequent NAD(+) depletion by oxidative stress may have consequent effects on inflammatory and stress responses as well as cellularsenescence. SIRT1 has been shown to undergo covalent oxidative modifications by cigarette smoke-derived oxidants/aldehydes, leading to posttranslational modifications, inactivation, and protein degradation. Furthermore, oxidant/carbonyl stress-mediated reduction of SIRT1 leads to the loss of its control on acetylation of target proteins including p53, RelA/p65, and FOXO3, thereby enhancing the inflammatory, prosenescent, and apoptotic responses, as well as endothelial dysfunction. In this review, the mechanisms of cigarette smoke/oxidant-mediated redox posttranslational modifications of SIRT1 and its roles in PARP1 and NF-κB activation, and FOXO3 and eNOS regulation, as well as chromatin remodeling/histone modifications during inflammaging, are discussed. Furthermore, we have also discussed various novel ways to activate SIRT1 either directly or indirectly, which may have therapeutic potential in attenuating inflammation and premature senescence involved in chronic lung diseases.

Endothelial senescence may contribute to the pathogenesis of age-related vascular disorders. Furthermore, chronic exposure to risk factors for cardiovascular disease (CVD) accelerates the effects of chronological aging by generating stress-dependent damages, including oxidative stress, therefore promoting stress-induced premature senescence. Our objective was to determine whether a chronic treatment with an antioxidant (N-acetyl-cystein, NAC) could delay senescence of endothelial cells (EC) isolated and cultured from arterial segments of patients with severe coronary artery disease. If EC were considered as one population (n=26), chronic NAC treatment slightly shortened telomere attrition rate associated with senescence but did not significantly delay the onset of endothelial senescence. However, in a subgroup of NAC-treated EC (n=15) cellularsenescence was significantly delayed, NAC decreased lipid peroxidation (HNE), activated the catalytic subunit of telomerase (hTERT) and inhibited telomere attrition. In contrast, in another subgroup of EC (n=11) characterized by initial short telomeres, no effect of NAC on HNE and high levels of DNA damages, the antioxidant was not beneficial on senescence, suggesting an irreversible stress-dependent damage. In conclusion, chronic exposure to NAC can delay senescence of diseased EC via hTERT activation and transient telomere stabilization, unless oxidative stress-associated cell damage has become irreversible.

Cellularsenescence is of growing interest due to its role in tumour suppression and its contribution to organismic ageing. This cellular state can be reached by replicative loss of telomeres or certain stresses in cell culture and is characterized by the termination of cell division; however, the cells remain metabolically active. To identify metabolites that are characteristic for senescent cells, extracts of human embryonic lung fibroblast (WI-38 cell line) have been investigated with NMR spectroscopy. Three different types of senescence have been characterized: replicative senescence, DNA damage-induced senescence (etoposide treatment) and oncogene-induced senescence (hyperactive RAF kinase). The metabolite pattern allows (I) discrimination of senescent and control cells and (II) discrimination of the three senescence types. Senescent cells show an increased ratio of glycerophosphocholine to phosphocholine independent from the type of senescence. The increase in glycerophosphocholine implicates a key role of phospholipid metabolism in cellularsenescence. The observed changes in the choline metabolism are diametrically opposite to the well-known changes in choline metabolism of tumour cells. As tumours responding to chemotherapeutic agents show a "glycerophosphocholine-to-phosphocholine switch" i.e. an increase in glycerophosphocholine, our metabolic data suggests that these malignant cells enter a senescent state emphasizing the role of senescence in tumour suppression.

XFE progeroid syndrome, a disease of accelerated aging caused by deficiency in the DNA repair endonuclease XPF-ERCC1, is modeled by Ercc1 knockout and hypomorphic mice. Tissues and primary cells from these mice senesce prematurely, offering a unique opportunity to identify factors that regulate senescence and aging. We compared microRNA (miRNA) expression in Ercc1−/− primary mouse embryonic fibroblasts (MEFs) and wild-type (WT) MEFs in different growth conditions to identify miRNAs that drive cellularsenescence. Microarray analysis showed three differentially expressed miRNAs in passage 7 (P7) Ercc1−/− MEFs grown at 20% O2 compared to Ercc1−/− MEFs grown at 3% O2. Thirty-six differentially expressed miRNAs were identified in Ercc1−/− MEFs at P7 compared to early passage (P3) in 3% O2. Eight of these miRNAs (miR-449a, miR-455*, miR-128, miR-497, miR-543, miR-450b-3p, miR-872 and miR-10b) were similarly downregulated in the liver of progeroid Ercc1−/Δ and old WT mice compared to adult WT mice, a tissue that senesces with aging. Three miRNAs (miR-449a, miR-455* and miR-128) were also downregulated in Ercc1−/Δ and WT old mice kidneys compared to young WT mice. We also discovered that the miRNA expression regulator Dicer is significantly downregulated in tissues of old mice and late passage cells compared to young controls. Collectively these results support the conclusion that the miRNAs identified may play an important role in staving off cellularsenescence and their altered expression could be indicative of aging. PMID:23852002

Cellularsenescence, a process that imposes permanent proliferative arrest on cells in response to various stressors, has emerged as a potentially important contributor to aging and age-related disease, and it is an attractive target for therapeutic exploitation. A wealth of information about senescence in cultured cells has been acquired over the past half century; however, senescence in living organisms is poorly understood, largely because of technical limitations relating to the identification and characterization of senescent cells in tissues and organs. Furthermore, newly recognized beneficial signaling functions of senescence suggest that indiscriminately targeting senescent cells or modulating their secretome for anti-aging therapy may have negative consequences. Here we discuss current progress and challenges in understanding the stressors that induce senescence in vivo, the cell types that are prone to senesce, and the autocrine and paracrine properties of senescent cells in the contexts of aging and age-related diseases as well as disease therapy. PMID:26646499

Cellularsenescence, a process that imposes permanent proliferative arrest on cells in response to various stressors, has emerged as a potentially important contributor to aging and age-related disease, and it is an attractive target for therapeutic exploitation. A wealth of information about senescence in cultured cells has been acquired over the past half century; however, senescence in living organisms is poorly understood, largely because of technical limitations relating to the identification and characterization of senescent cells in tissues and organs. Furthermore, newly recognized beneficial signaling functions of senescence suggest that indiscriminately targeting senescent cells or modulating their secretome for anti-aging therapy may have negative consequences. Here we discuss current progress and challenges in understanding the stressors that induce senescence in vivo, the cell types that are prone to senesce, and the autocrine and paracrine properties of senescent cells in the contexts of aging and age-related diseases as well as disease therapy.

Holliday junction recognition protein (HJURP), a centromere protein-A (CENP-A) histone chaperone, mediates centromere-specific assembly of CENP-A nucleosome, contributing to high-fidelity chromosome segregation during cell division. However, the role of HJURP in cellularsenescence of human primary cells remains unclear. We found that the expression levels of HJURP decreased in human dermal fibroblasts and umbilical vein endothelial cells in replicative or premature senescence. Ectopic expression of HJURP in senescent cells partially overcame cell senescence. Conversely, downregulation of HJURP in young cells led to premature senescence. p53 knockdown, but not p16 knockdown, abolished senescence phenotypes caused by HJURP reduction. These data suggest that HJURP plays an important role in the regulation of cellularsenescence through a p53-dependent pathway and might contribute to tissue or organismal aging and protection of cellular transformation.

Senescenceaccelerated mice (SAM) are susceptible to developing vascular dysfunction and remodeling. Food intake and type of diet have also been identified as determining factors in vascular remodeling. However, the interplay between senescence and diet in vascular remodeling is largely unknown. We aimed to analyze structure of large (aorta) and small (mesenteric; MA) arteries from seven-month-old SAM prone (SAMP8) and resistant (SAMR1) mice that received a Western-type high-fat diet (WD; 8weeks). Aortic structure was assessed by morphometric analysis of hematoxylin and eosin-stained cross sections, and collagen content by qRT-PCR, immunofluorescence and picrosirius red. In MAs, structural and mechanical properties were measured by pressure myography; elastin and collagen content by qRT-PCR and immunofluorescence; nuclei distribution by confocal microscopy; and apoptosis by qRT-PCR and TUNEL assay. In aorta, wall thickness (WT), but not cross-sectional area (CSA), was increased by senescence, and WD only increased WT in SAMR1. WD intake, but not senescence, was associated with increased collagen deposition. In MAs, senescence diminished WT and CSA, without altering collagen and elastin deposition, reduced the number of MA wall cells, and increased pro apoptotic activation. WD consumption promoted in SAMR1 the same remodeling observed with senescence, while in SAMP8 the senescence-associated changes remained unaffected. The mechanisms involved in WD-induced MA remodeling in SAMR1 mimicked those observed in senescence per se. Our study reveals qualitatively different remodeling in aortas and MAs from senescent mice. Consumption of a WD induced remodeling of the SAMR1 vasculature similar to that induced by senescence, while it did not promote any further alteration in the latter. Therefore, we propose that increased consumption of fat-enriched diets could promote acceleratedsenescence of the non-senescent vasculature, although it does not exacerbate vascular

Mesenchymal stem cells (MSCs) are multipotent progenitor cells that represent a promising approach in the field of regenerative medicine; however, this potential diminishes with senescence. Pigment epithelium-derived factor (PEDF) gives some protection by reducing oxidative stress, which is known to acceleratecellularsenescence. Thus we hypothesized that PEDF could delay senescence during MSC expansion by reducing oxidative stress. Proliferation and differentiation potentials, oxidative stress, senescence and p53/p16 expressions have been examined. In MSCs cultured under normoxic conditions treated with PEDF, proliferative lifespan in vitro was significantly increased compared with control group not given PEDF, with ∼10 additional population doublings (PD) occurring before terminal growth arrest. Most of the MSCs cultured under normoxic conditions ceased to proliferate after 20-28 PD, while few senescent cells were found in the hypoxic, PEDF-hypoxic and PEDF-normoxic cultures; this was associated with downregulation of p53 and p16 expression and decreased oxidative stress. PEDF also preserved differentiation potentials of MSCs compared with the control group. Thus PEDF suppression of oxidative stress delays cellularsenescence and allows greater expansion of MSCs.

Cellularsenescence, a form of stable cell cycle arrest that is traditionally associated with tumour suppression, has been recently found to occur during mammalian development. Here, we show that cell senescence is an intrinsic part of the developmental programme in amphibians. Programmed senescence occurs in specific structures during defined time windows during amphibian development. It contributes to the physiological degeneration of the amphibian pronephros and to the development of the cement gland and oral cavity. In both contexts, senescence depends on TGFβ but is independent of ERK/MAPK activation. Furthermore, elimination of senescent cells through temporary TGFβ inhibition leads to developmental defects. Our findings uncover conserved and new roles of senescence in vertebrate organogenesis and support the view that cellularsenescence may have arisen in evolution as a developmental mechanism. PMID:27888193

Cell senescence is associated with the secretion of many factors, the so-called “senescence-associated secretory phenotype”, which may alter tissue microenvironment, stimulating the organism to clean up senescent cells and replace them with newly divided ones. Therefore, although no longer dividing, these cells are still metabolically active and influence the surrounding tissue. Much attention has been recently focused not only on soluble factors released by senescent cells, but also on extracellular vesicles as conveyors of senescence signals outside the cell. Here, we give an overview of the role of extracellular vesicles in biological processes and signaling pathways related to senescence and aging. PMID:27571072

Accidental high-dose radiation exposures can lead to multi-organ injuries, including radiation dermatitis. The types of cellular damage leading to radiation dermatitis are not completely understood. To identify the cellular mechanisms that underlie radiation-induced skin injury in vivo, we evaluated the time-course of cellular effects of radiation (14, 16 or 17 Gy X-rays; 0.5 Gy/min) in the skin of C57BL/6 mice. Irradiation of 14 Gy induced mild inflammation, observed histologically, but no visible hair loss or erythema. However, 16 or 17 Gy radiation induced dry desquamation, erythema and mild ulceration, detectable within 14 days post-irradiation. Histological evaluation revealed inflammation with mast cell infiltration within 14 days. Fibrosis occurred 80 days following 17 Gy irradiation, with collagen deposition, admixed with neutrophilic dermatitis, and necrotic debris. We found that in cultures of normal human keratinocytes, exposure to 17.9 Gy irradiation caused the upregulation of p21/waf1, a marker of senescence. Using western blot analysis of 17.9 Gy-irradiated mice skin samples, we also detected a marker of acceleratedsenescence (p21/waf1) 7 days post-irradiation, and a marker of cellular apoptosis (activated caspase-3) at 30 days, both preceding histological evidence of inflammatory infiltrates. Immunohistochemistry revealed reduced epithelial stem cells from hair follicles 14-30 days post-irradiation. Furthermore, p21/waf1 expression was increased in the region of the hair follicle stem cells at 14 days post 17 Gy irradiation. These data indicate that radiation induces acceleratedcellularsenescence in the region of the stem cell population of the skin.

Doxorubicin, an anthracycline antibiotic, is a commonly used anticancer drug. In spite of its widespread usage, its therapeutic effect is limited by its cardiotoxicity. On the cellular level, Doxorubicin-induced cardiotoxicity manifests as stress induced premature senescence. Previously, we demonstrated that plasminogen activator inhibitor-1 (PAI-1), a potent inhibitor of serine proteases, is an important biomarker and regulator of cellularsenescence and aging. Here, we tested the hypothesis that pharmacological inhibition of cellular PAI-1 protects against stress- and aging-induced cellularsenescence and delineated the molecular basis of protective action of PAI-1 inhibition. Results show that TM5441, a potent small molecule inhibitor of PAI-1, effectively prevents Doxorubicin-induced senescence in cardiomyocytes, fibroblasts and endothelial cells. TM5441 exerts its inhibitory effect on Doxorubicin-induced cellularsenescence by decreasing reactive oxygen species generation, induction of antioxidants like catalase and suppression of stress-induced senescence cadre p53, p21, p16, PAI-1 and IGFBP3. Importantly, TM5441 also reduces replicative senescence of fibroblasts. Together these results for the first time demonstrate the efficacy of PAI-1 inhibitor in prevention of Doxorubicin-induced and replicative senescence in normal cells. Thus PAI-1 inhibitor may form an important adjuvant component of chemotherapy regimens, limiting not only Doxorubicin-induced cardiac senescence but also ameliorating the prothrombotic profile. PMID:27736799

Although senescence is a defining property of euploid mammalian cells, its physiologic basis remains obscure. Previously, cell kinetics properties of normal tissue cells have not been considered in models for senescence. We now provide evidence that senescence is in fact the natural consequence of normal in vivo somatic stem cell kinetics extended in culture. This concept of senescence is based on our discovery that cells engineered to conditionally express the well-recognized tumor suppressor protein and senescence factor, p53, exhibit asymmetric cell kinetics. In vivo, asymmetric cell kinetics are essential for maintenance of somatic stem cells; ex vivo, the same cell kinetics yield senescence as a simple kinetic endpoint. This new “asymmetric cell kinetics model” for senescence suggests novel strategies for the isolation and propagation of somatic tissue stem cells in culture. PMID:12488624

Background Progressive neurological dysfunction is a key aspect of human aging. Because of underlying differences in the aging of mice and humans, useful mouse models have been difficult to obtain and study. We have used gene-expression analysis and polymorphism screening to study molecular senescence of the retina and hippocampus in two rare inbred mouse models of accelerated neurological senescence (SAMP8 and SAMP10) that closely mimic human neurological aging, and in a related normal strain (SAMR1) and an unrelated normal strain (C57BL/6J). Results The majority of age-related gene expression changes were strain-specific, with only a few common pathways found for normal and accelerated neurological aging. Polymorphism screening led to the identification of mutations that could have a direct impact on important disease processes, including a mutation in a fibroblast growth factor gene, Fgf1, and a mutation in and ectopic expression of the gene for the chemokine CCL19, which is involved in the inflammatory response. Conclusion We show that combining the study of inbred mouse strains with interesting traits and gene-expression profiling can lead to the discovery of genes important for complex phenotypes. Furthermore, full-genome polymorphism detection, sequencing and gene-expression profiling of inbred mouse strains with interesting phenotypic differences may provide unique insights into the molecular genetics of late-manifesting complex diseases. PMID:15960800

In this work, it was examined the possibility of lipopolysaccharide (LPS) causing cellularsenescence in lung alveolar epithelial cells. Then, it was clarified how this cellularsenescence phenomenon is associated with oxidative stress effect induced by LPS and whether antioxidants could inhibit reduced cellular viability by oxidant stress effect of LPS. In cell viability using cell counting kit-8, exposure to LPS decreased cellular viability and induced growth arrest in a concentration-dependent manner. The pre-apoptotic concentration of LPS was determined by caspase activation using a Caspase-Glo 3/7 luminescence assay kit. This concentration of LPS caused morphologic characteristics shown in senescent cells and elevated senescence-associated β-galactosidase activity. In addition, lysosomal content associated with senescence was increased by LPS at the pre-apoptotic concentration. However, this concentration of LPS did not shorten the telomere length. Exposure to LPS resulted in the formation of hydrogen peroxide in a concentration-dependent manner. The ability of LPS to reduce cellular viability was inhibited by the presence of glutathione. This study revealed that LPS could induce cellularsenescence in lung alveloar epithelial cells, and these phenomena were closely associated with hydrogen peroxide production by LPS. Taken together, it is suggested that LPS-induced cellularsenescence may play an important role in limiting the tissue repair response after sepsis.

Highlights: Black-Right-Pointing-Pointer Cellularsenescence plays an important role in tumorigenesis and aging process. Black-Right-Pointing-Pointer We demonstrated IGF-I enhanced cellularsenescence in primary confluent cells. Black-Right-Pointing-Pointer IGF-I enhanced cellularsenescence in the ROS and p53-dependent manner. Black-Right-Pointing-Pointer These results may explain the underlying mechanisms of IGF-I involvement in tumorigenesis and in regulation of aging. -- Abstract: Cellularsenescence is characterized by growth arrest, enlarged and flattened cell morphology, the expression of senescence-associated {beta}-galactosidase (SA-{beta}-gal), and by activation of tumor suppressor networks. Insulin-like growth factor-I (IGF-I) plays a critical role in cellular growth, proliferation, tumorigenesis, and regulation of aging. In the present study, we show that IGF-I enhances cellularsenescence in mouse, rat, and human primary cells in the confluent state. IGF-I induced expression of a DNA damage marker, {gamma}H2AX, the increased levels of p53 and p21 proteins, and activated SA-{beta}-gal. In the confluent state, an altered downstream signaling of IGF-I receptor was observed. Treatment with a reactive oxygen species (ROS) scavenger, N-acetylcystein (NAC) significantly suppressed induction of these markers, indicating that ROS are involved in the induction of cellularsenescence by IGF-I. In p53-null mouse embryonic fibroblasts, the IGF-I-induced augmentation of SA-{beta}-gal and p21 was inhibited, demonstrating that p53 is required for cellularsenescence induced by IGF-I. Thus, these data reveal a novel pathway whereby IGF-I enhances cellularsenescence in the ROS and p53-dependent manner and may explain the underlying mechanisms of IGF-I involvement in tumorigenesis and in regulation of aging.

This study investigated the effect of isoflavone aglycone (IA) on the learning and memory performance of senescence-accelerated mice, and explored its neural protective mechanism. Results showed that SAM-P/8 senescence-accelerated mice treated with IA performed significantly better in the Y-maze cognitive test than the no treatment control (P less…

Dental stem cells such as human dental follicle cells (DFCs) have opened new promising treatment alternatives for today's dental health issues such as periodontal tissue regeneration. However, cellularsenescence represents a restricting factor to cultured stem cells, resulting in limited lifespan and reduced cell differentiation potential. Therefore, this study evaluated if and how DFCs exhibit features of cellularsenescence after being expanded in cell culture. The cell proliferation of DFCs decreased, while the cell size increased during prolonged cell culture. Moreover, DFCs expressed the senescence-associated β-galactosidase after a prolonged cell culture. The onset of senescence inhibited both the induction of osteoblast markers RUNX2 and osteopontin and the biomineralization of DFCs after stimulation of the osteogenic differentiation. In conclusion, we showed that a prolonged cell culture induces cellularsenescence and inhibits the osteogenic differentiation in DFCs.

Cystatin SN (CST1), a known inhibitor of cathepsin B (CatB), has important roles in tumor development. Paradoxically, CatB is a member of the cysteine cathepsin family that acts in cellular processes, such as tumor development and invasion. However, the relationship between CST1 and CatB, and their roles in tumor development are poorly understood. In this study, we observed that the knockdown of CST1 induced the activity of senescence-associated β-galactosidase, a marker of cellularsenescence, and expression of senescence-associated secretory phenotype genes, including interleukin-6 and chemokine (C-C motif) ligand 20, in MDA-MB-231 and SW480 cancer cells. Furthermore, CST1 knockdown decreased extracellular CatB activity, and direct CatB inhibition, using specific inhibitors or shCatB, induced cellularsenescence. Reconstitution of CST1 restored CatB activity and inhibited cellularsenescence in CST1 knockdown cells. CST1 knockdown or CatB inhibition increased glycogen synthase (GS) kinase 3β phosphorylation at serine 9, resulting in the activation of GS and the induction of glycogen accumulation associated with cellularsenescence. Importantly, CST1 knockdown suppressed cancer cell proliferation, soft agar colony growth and tumor growth in a xenograft model. These results indicate that CST1-mediated extracellular CatB activity enhances tumor development by preventing cellularsenescence. Our findings suggest that antagonists of CST1 or inhibitors of CatB are potential anticancer agents.

During leaf senescence, degradation of chloroplasts precede to changes in nuclei and other cytoplasmic organelles, RuBisCO stability is progressively lost, grana lose their structure, plastidial DNA becomes distorted and degraded, the number of plastoglobuli increases and abundant senescence-associated vesicles containing electronically dense particles emerge from chloroplasts pouring their content into the central vacuole. This study examines quinoa leaf tissues during development and senescence using a range of well-established markers of programmed cell death (PCD), including: morphological changes in nuclei and chloroplasts, degradation of RuBisCO, changes in chlorophyll content, DNA degradation, variations in ploidy levels, and changes in nuclease profiles. TUNEL reaction and DNA electrophoresis demonstrated that DNA fragmentation in nuclei occurs at early senescence, which correlates with induction of specific nucleases. During senescence, metabolic activity is high and nuclei endoreduplicate, peaking at 4C. At this time, TEM images showed some healthy nuclei with condensed chromatin and nucleoli. We have found that DNA fragmentation, induction of senescence-associated nucleases and endoreduplication take place during leaf senescence. This provides a starting point for further research aiming to identify key genes involved in the senescence of quinoa leaves.

Notch activity regulates tumor biology in a context-dependent and complex manner. Notch may act as an oncogene or a tumor-suppressor gene even within the same tumor type. Recently, Notch signaling has been implicated in cellularsenescence. Yet, it remains unclear as to how cellularsenescence checkpoint functions may interact with Notch-mediated oncogenic and tumor-suppressor activities. Herein, we used genetically engineered human esophageal keratinocytes and esophageal squamous cell carcinoma cells to delineate the functional consequences of Notch activation and inhibition along with pharmacological intervention and RNA interference experiments. When expressed in a tetracycline-inducible manner, the ectopically expressed activated form of Notch1 (ICN1) displayed oncogene-like characteristics inducing cellularsenescence corroborated by the induction of G0/G1 cell-cycle arrest, Rb dephosphorylation, flat and enlarged cell morphology and senescence-associated β-galactosidase activity. Notch-induced senescence involves canonical CSL/RBPJ-dependent transcriptional activity and the p16(INK4A)-Rb pathway. Loss of p16(INK4A) or the presence of human papilloma virus (HPV) E6/E7 oncogene products not only prevented ICN1 from inducing senescence but permitted ICN1 to facilitate anchorage-independent colony formation and xenograft tumor growth with increased cell proliferation and reduced squamous-cell differentiation. Moreover, Notch1 appears to mediate replicative senescence as well as transforming growth factor-β-induced cellularsenescence in non-transformed cells and that HPV E6/E7 targets Notch1 for inactivation to prevent senescence, revealing a tumor-suppressor attribute of endogenous Notch1. In aggregate, cellularsenescence checkpoint functions may influence dichotomous Notch activities in the neoplastic context.

MicroRNA-194 (miR-194), a typical p53 responsive miRNA, serves as a tumor suppressor similar as p53, and has been demonstrated to play an anti-proliferation role in various human cancers. In spite of the pivotal role of p53 during aging process, the knowledge of miR-194's contribution to cellularsenescence is limited. We herein sought to explore the role of miR-194 in the replicative senescence and stress-induced senescence of mouse embryonic fibroblasts. Our results unraveled that, compared to young cells, miR-194 is highly expressed in senescent cells, and extra expression of miR-194 significantly triggers the replicative senescence of MEFs and H2 O2 -induced senescence of NIH/3T3 cells, while inhibition of miR-194 exhibited the opposite effect. We further unveiled that DNMT3A was a direct and authentic target of miR-194, which has been reported to be closely associated with cellularsenescence. Taken together, our data suggest that miR-194 may significantly promote the development of cellularsenescence in mouse embryonic fibroblasts, which potentially occurs through inhibiting the DNMT3A expression.

Polo-like kinase 1 (PLK1) plays a key role in various stages of mitosis from entry into M phase to exit from mitosis. However, its role in cellularsenescence remains to be determined. Therefore, the effects of PLK1 on cellularsenescence in human primary cells were investigated. We found that expression of PLK1 decreased in human dermal fibroblasts and human umbilical vein endothelial cells under replicative senescence and premature senescence induced by adriamycin. PLK1 knockdown with PLK1 small interfering RNAs in young cells induced premature senescence. In contrast, upregulation of PLK1 in old cells partially reversed senescence phenotypes. Cellularsenescence by PLK1 inhibition was observed in p16 knockdown cells but not in p53 knockdown cells. Our data suggest that PLK1 repression might result in cellularsenescence in human primary cells via a p53-dependent pathway.

Cellularsenescence has been recently shown to play an important role in opposing tumour initiation and promotion. Senescence induced by oncogenes or loss of tumour suppressor genes is thought to critically dependent on the induction of the p19Arf-p53 pathway. The Skp2 E3-ubiquitin ligase can act as a proto-oncogene and its aberrant overexpression is frequently observed in human cancers. Here we show that although Skp2 inactivation on its own does not induce cellularsenescence, aberrant proto-oncogenic signals as well as inactivation of tumour suppressor genes do trigger a potent, tumor-suppressive senescence response in mice and cells devoid of Skp2. Notably, Skp2 inactivation and oncogenic stress driven senescence neither elicits activation of the p19Arf-p53 pathway nor DNA damage, but instead depends on ATF4, p27, and p21. We further demonstrate that genetic Skp2 inactivation evokes cellularsenescence even in oncogenic conditions in which the p19Arf/p53 response is impaired, whereas a Skp2-SCF complex inhibitor can trigger cellularsenescence in p53/PTEN deficient cells and tumour regression in preclinical studies. Our findings therefore provide proof of principle evidence that Skp2 pharmacological inhibition may represent a general approach for cancer prevention and therapy. PMID:20237562

Liver fibrosis occurs as a wound-healing response to chronic hepatic injuries irrespective of the underlying etiology and may progress to life-threatening cirrhosis. Here we show that CCN1, a matricellular protein of the CCN (CYR61/CTGF/NOV) family, is accumulated in hepatocytes of human cirrhotic livers. CCN1 is not required for liver development or regeneration, since these processes are normal in mice with hepatocyte-specific Ccn1 deletion. However, Ccn1 expression is upregulated upon liver injuries and functions to inhibit liver fibrogenesis induced by either carbon tetrachloride intoxication or bile duct ligation and promote fibrosis regression. CCN1 acts by triggering cellularsenescence in activated hepatic stellate cells and portal fibroblasts by engaging integrin α6β1 to induce reactive oxygen species accumulation through the RAC1-NADPH oxidase 1 enzyme complex, whereupon the senescent cells express an antifibrosis genetic program. Mice with hepatocyte-specific Ccn1 deletion suffer exacerbated fibrosis with a concomitant deficit in cellularsenescence, whereas overexpression of hepatic Ccn1 reduces liver fibrosis with enhanced senescence. Furthermore, tail vein delivery of purified CCN1 protein accelerates fibrosis regression in mice with established fibrosis. These findings reveal a novel integrin-dependent mechanism of fibrosis resolution in chronic liver injury and identify the CCN1 signaling pathway as a potential target for therapeutic intervention.

While cellularsenescence is a critical mechanism to prevent malignant transformation of potentially mutated cells, persistence of senescent cells can also promote cancer and aging phenotypes. NonO/p54nrb and PSF are multifunctional hnRNPs typically found as a complex exclusively within the nuclei of all mammalian cells. We demonstrate here that either increase or reduction of expression of either factor results in cellularsenescence. Coincident with this, we observe expulsion of NonO and PSF-containing nuclear paraspeckles and posttranslational modification at G2/M. That senescence is mediated most robustly by overexpression of a cytoplasmic C-truncated form of NonO further indicated that translocation of NonO and PSF from the nucleus is critical to senescence induction. Modulation of NonO and PSF expression just prior to or coincident with senescence induction disrupts the normally heterodimeric NonO-PSF nuclear complex resulting in a dramatic shift in stoichiometry to heterotetramers and monomer with highest accumulation within the cytoplasm. This is accompanied by prototypic cell cycle checkpoint activation and chromatin condensation. These observations identify yet another role for these multifunctional factors and provide a hitherto unprecedented mechanism for cellularsenescence and nuclear-cytoplasmic trafficking. PMID:27992859

While cellularsenescence is a critical mechanism to prevent malignant transformation of potentially mutated cells, persistence of senescent cells can also promote cancer and aging phenotypes. NonO/p54nrb and PSF are multifunctional hnRNPs typically found as a complex exclusively within the nuclei of all mammalian cells. We demonstrate here that either increase or reduction of expression of either factor results in cellularsenescence. Coincident with this, we observe expulsion of NonO and PSF-containing nuclear paraspeckles and posttranslational modification at G2/M. That senescence is mediated most robustly by overexpression of a cytoplasmic C-truncated form of NonO further indicated that translocation of NonO and PSF from the nucleus is critical to senescence induction. Modulation of NonO and PSF expression just prior to or coincident with senescence induction disrupts the normally heterodimeric NonO-PSF nuclear complex resulting in a dramatic shift in stoichiometry to heterotetramers and monomer with highest accumulation within the cytoplasm. This is accompanied by prototypic cell cycle checkpoint activation and chromatin condensation. These observations identify yet another role for these multifunctional factors and provide a hitherto unprecedented mechanism for cellularsenescence and nuclear-cytoplasmic trafficking.

Oncogene-induced senescence is thought to act as a barrier to tumorigenesis by arresting cells at risk of malignant transformation. Nevertheless, numerous findings suggest that senescent cells may conversely promote tumor progression through the development of the senescence-associated secretome they produce. It is likely that the composition and the physiological consequences mediated by the senescence secretome are dependent on the oncogenes that trigger the senescence program. Breast cancer represents a heterogenous disease that can be divided into breast cancer subtypes due to different subsets of genetic and epigenetic abnormalities. As tumor initiation and progression of these breast cancer subtypes is triggered by diverse oncogenic stimuli, differences in the senescence secretomes within breast tumors might be responsible for tumor initiation, progression, metastasis and therapeutic response. Many studies have addressed the role of senescence as a barrier to tumor progression using murine xenograft models. However, few investigations have been performed to elucidate the degree to which senescent tumor cells are present within untreated human tumors, and if present, whether these senescent tumor cells may play a role in disease progression. In the present study we analysed the appearance of senescent cells within invasive breast cancers. Detection of cellularsenescence by the use of SAβ-galactosidase (SAβ-gal) staining within invasive breast carcinoms from 129 untreated patients revealed differences in the amount of SAβ-gal+ tumor cells between breast cancer subtypes. The highest percentages of SAβ-gal+ tumor cells were found in HER2-positive and luminal A breast carcinomas whereas triple negative tumors showed either little or no positivity. PMID:27713152

A compensatory response, viz. in vivo recovery from radiation damage to mitochondria, occurs in preclimacteric pear fruits (Pyrus communis L.) treated with ionizing radiation. The compensatory response is absent or markedly impaired in senescent fruits irradiated at or near the climacteric peak. Senescent cells failed to recover from harmful effects of radiation on: 1) mitochondrial yield, 2) in vivo incorporation of amino acids into mitochondrial protein, and 3) mitochondrial respiratory control and ADP/O. A diminished response to “split-dose” irradiation and a delayed rate of recovery confirmed the degeneracy and loss of compensatory power with cell age. A loss of restorative activity, especially in mitochondria that supply the cell with essential energy, may underlie the more obvious signs of cumulative stress that accompany cellularsenescence. Use of ionizing radiation as an investigative tool and the molecular implications of radiation damage, recovery, and cellularsenescence are discussed. PMID:16656887

Moxidectin has been used safely as an antiparasitic in many animal species, including for the eradication of the mouse fur mite, Mycoptes musculinus. Although no side effects of moxidectin have previously been reported to occur in mice, 2 strains of the senescence-accelerated mouse (SAMP8 and SAMR1) sustained considerable mortality after routine prophylactic treatment. To investigate the mechanism underlying this effect, moxidectin toxicosis in these mice was evaluated in a controlled study. Moxidectin was applied topically (0.015 mg), and drug concentrations in both brain and serum were analyzed by using HPLC coupled with mass spectrometry. The moxidectin concentration in brain of SAMP8 mice was 18 times that in controls, and that in brain of SAMR1 mice was 14 times higher than in controls, whereas serum moxidectin concentrations did not differ significantly among the 3 strains. Because deficiency of the blood-brain barrier protein P-glycoprotein leads to sensitivity to this class of drugs in other SAM mice, Pgp immunohistochemistry of brain sections from a subset of mice was performed to determine whether this commercially available analysis could predict sensitivity to this class of drug. The staining analysis showed no difference among the strains of mice, indicating that this test does not correlate with sensitivity. In addition, no gross or histologic evidence of organ toxicity was found in brain, liver, lung, or kidney. This report shows that topically applied moxidectin at a standard dose accumulates in the CNS causing toxicosis in both SAMP8 and SAMR1 mice.

During an investigation into the use of oil emulsions in foliar sprays, it was found that silicone oil emulsions accelerated the senescence of the primary leaves of bean (Phaseolus vulgaris) plants. It was shown that acceleratedsenescence was not a result of the reduced transpiration rates found in silicone-sprayed leaves. Furthermore, the silicone oil emulsions did not induce leakiness in plant cell membranes. The senescence-enhancing effect seems to be connected with the ability of the silicone oil emulsions to penetrate into the leaf interior.

The DNA damage response (DDR) orchestrates DNA repair and halts cell cycle. If damage is not resolved, cells can enter into an irreversible state of proliferative arrest called cellularsenescence. Organismal ageing in mammals is associated with accumulation of markers of cellularsenescence and DDR persistence at telomeres. Since the vast majority of the cells in mammals are non-proliferating, how do they age? Are telomeres involved? Also oncogene activation causes cellularsenescence due to altered DNA replication and DDR activation in particular at the telomeres. Is there a common mechanism shared among apparently distinct types of cellularsenescence? And what is the role of telomeric DNA damage?

Described is an in vitro model of premature senescence in pulmonary adenocarcinoma A549 cells induced by persistent DNA replication stress in response to treatment with the DNA damaging drug mitoxantrone (Mxt). The degree of cellularsenescence, based on characteristic changes in cell morphology, is measured by laser scanning cytometry. Specifically, the flattening of cells grown on slides (considered the hallmark of cellularsenescence) is measured as the decline in local intensity of DNA-associated DAPI fluorescence (represented by maximal pixels). This change is paralleled by an increase in nuclear area. Thus, the ratio of mean intensity of maximal pixels to nuclear area provides a very sensitive morphometric biomarker for the degree of senescence. This analysis is combined with immunocytochemical detection of senescence markers, such as overexpression of cyclin kinase inhibitors (e.g., p21(WAF1) ) and phosphorylation of ribosomal protein S6 (rpS6), a key marker associated with aging/senescence that is detected using a phospho-specific antibody. These biomarker indices are presented in quantitative terms defined as a senescence index (SI), which is the fraction of the marker in test cultures relative to the same marker in exponentially growing control cultures. This system can be used to evaluate the anti-aging potential of test agents by assessing attenuation of maximal senescence. As an example, the inclusion of berberine, a natural alkaloid with reported anti-aging properties and a long history of use in traditional Chinese medicine, is shown to markedly attenuate the Mxt-induced SI and phosphorylation of rpS6. The multivariate analysis of senescence markers by laser scanning cytometry offers a promising tool to explore the potential anti-aging properties of a variety agents.

Described is an in vitro model of premature senescence in pulmonary adenocarcinoma A549 cells induced by persistent DNA replication stress in response to treatment with the DNA damaging drug mitoxantrone (Mxt). The degree of cellularsenescence, based on characteristic changes in cell morphology, is measured by laser scanning cytometry. Specifically, the flattening of cells grown on slides (considered the hallmark of cellularsenescence) is measured as the decline in local intensity of DNA-associated DAPI fluorescence (represented by maximal pixels). This change is paralleled by an increase in nuclear area. Thus, the ratio of mean intensity of maximal pixels to nuclear area provides a very sensitive morphometric biomarker for the degree of senescence. This analysis is combined with immunocytochemical detection of senescence markers, such as overexpression of cyclin kinase inhibitors (e.g., p21WAF1) and phosphorylation of ribosomal protein S6 (rpS6), a key marker associated with aging/senescence that is detected using a phospho-specific antibody. These biomarker indices are presented in quantitative terms defined as a senescence index (SI), which is the fraction of the marker in test cultures relative to the same marker in exponentially growing control cultures. This system can be used to evaluate the anti-aging potential of test agents by assessing attenuation of maximal senescence. As an example, the inclusion of berberine, a natural alkaloid with reported anti-aging properties and a long history of use in traditional Chinese medicine, is shown to markedly attenuate the Mxt-induced SI and phosphorylation of rpS6. The multivariate analysis of senescence markers by laser scanning cytometry offers a promising tool to explore the potential anti-aging properties of a variety agents. PMID:24984966

Although pituitary adenomas represent 25% of intracranial tumors, they are usually benign, with the mechanisms by which these tumors usually avoid an invasive profile and metastatic growth development still remaining unclear. In this context, cellularsenescence might constitute a plausible explanation for the benign nature of pituitary adenomas. In this study, we investigated the emergence of cellularsenescence as a growth control mechanism during the progression of estrogen-induced pituitary tumors. The quantification of Ki67-immunopositive cells in the pituitaries of estrogenized male rats after 10, 20, 40, and 60 days revealed that the mitogenic potential rate was not sustained for the whole period analyzed and successively decreased after 10 days of estrogen exposure. In addition, the expression of cellularsenescence features, such as the progressive rise in the enzymatic senescence-associated b-galactosidase (SA-b-gal) activity, IL6, IL1b, and TGFb expression, was observed throughout pituitary tumor development. Furthermore, tumoral pituitary cells also displayed nuclear pATM expression, indicating activated DNA damage signaling, with a significant increase in p21 expression also being detected. The associations among DNA damage signaling activation, SA-b-gal expression, and p21 may provide a reliable combination of senescence-associated markers for in vivo pituitary senescence detection. These results suggest a role for this cellular process in the regulation of pituitary cell growth. Thus, cellularsenescence should be conceived as a contributing component to the benign nature of pituitary adenomas, thereby influencing the capability of the pituitary gland to avoid unregulated cell proliferation.

LMNA mutations are responsible for a variety of genetic disorders, including muscular dystrophy, lipodystrophy, and certain progeroid syndromes, notably Hutchinson-Gilford Progeria. Although a number of clinical features of these disorders are suggestive of accelerated aging, it is not known whether cells derived from these patients exhibit cellular phenotypes associated with accelerated aging. We examined a series of isogenic skin fibroblast lines transfected with LMNA constructs bearing known pathogenic point mutations or deletion mutations found in progeroid syndromes. Fibroblasts overexpressing mutant lamin A exhibited accelerated rates of loss of telomeres and shortened replicative lifespans, in addition to abnormal nuclear morphology. To our surprise, these abnormalities were also observed in lines overexpressing wild-type lamin A. Copy number variants are common in human populations; those involving LMNA, whether arising meiotically or mitotically, might lead to progeroid phenotypes. In an initial pilot study of 23 progeroid cases without detectible WRN or LMNA mutations, however, no cases of altered LMNA copy number were detected. Nevertheless, our findings raise a hypothesis that changes in lamina organization may cause accelerated telomere attrition, with different kinetics for overexpession of wild-type and mutant lamin A, which leads to rapid replicative senescence and progroid phenotypes. PMID:17870066

Cellularsenescence, a state of essentially irreversible proliferation arrest, serves as a potent tumour suppressor mechanism. However, accumulation of senescent cells with chronological age is likely to contribute to loss of tissue and organ function and organismal aging. A crucial biochemical modulator of aging is mTOR; here, we have addressed the question of whether acute mTORC inhibition in near-senescent cells can modify phenotypes of senescence. We show that acute short term treatment of human skin fibroblasts with low dose ATP mimetic pan-mTORC inhibitor AZD8055 leads to reversal of many phenotypes that develop as cells near replicative senescence, including reduction in cell size and granularity, loss of SA-β-gal staining and reacquisition of fibroblastic spindle morphology. AZD8055 treatment also induced rearrangement of the actin cytoskeleton, providing a possible mechanism of action for the observed rejuvenation. Importantly, short-term drug exposure had no detrimental effects on cell proliferation control across the life-course of the fibroblasts. Our findings suggest that combined inhibition of both mTORC1 and mTORC2 may provide a promising strategy to reverse the development of senescence-associated features in near-senescent cells. PMID:26851731

A novel target of NESH-SH3 (TARSH) was identified as a cellularsenescence related gene in mouse embryonic fibroblasts (MEFs) replicative senescence, the expression of which has been suppressed in primary clinical lung cancer specimens. However, the molecular mechanism underlying the regulation of TARSH involved in pulmonary tumorigenesis remains unclear. Here we demonstrate that the reduction of TARSH gene expression by short hairpin RNA (shRNA) system robustly inhibited the MEFs proliferation with increase in senescence-associated {beta}-galactosidase (SA-{beta}-gal) activity. Using p53{sup -/-} MEFs, we further suggest that this growth arrest by loss of TARSH is evoked by p53-dependent p21{sup Cip1} accumulation. Moreover, we also reveal that TARSH reduction induces multicentrosome in MEFs, which is linked in chromosome instability and tumor development. These results suggest that TARSH plays an important role in proliferation of replicative senescence and may serve as a trigger of tumor development.

Taurine (2-aminoethanesulfonic acid) is found in milimolar concentrations in mammalian tissues. One of its main functions is osmoregulation; however, it also exhibits cytoprotective activity by diminishing injury caused by stress and disease. Taurine depletion is associated with several defects, many of which are found in the aging animal, suggesting that taurine might exert anti-aging actions. Therefore, in the present study, we examined the hypothesis that taurine depletion accelerates aging by reducing longevity and accelerating aging-associated tissue damage. Tissue taurine depletion in taurine transporter knockout (TauTKO) mouse was found to shorten lifespan and accelerate skeletal muscle histological and functional defects, including an increase in central nuclei containing myotubes, a reduction in mitochondrial complex 1 activity and an induction in an aging biomarker, Cyclin-dependent kinase 4 inhibitor A (p16INK4a). Tissue taurine depletion also enhances unfolded protein response (UPR), which may be associated with an improvement in protein folding by taurine. Our data reveal that tissue taurine depletion affects longevity and cellularsenescence; an effect possibly linked to a disturbance in protein folding.

Cellularsenescence is a stable state of proliferative arrest that provides a barrier to malignant transformation and contributes to the antitumor activity of certain chemotherapies. Senescent cells can accumulate senescence-associated heterochromatic foci (SAHFs), which may provide a chromatin buffer that prevents activation of proliferation-associated genes by mitogenic transcription factors. Surprisingly, we show that the High-Mobility Group A (HMGA) proteins, which can promote tumorigenesis, accumulate on the chromatin of senescent fibroblasts and are essential structural components of SAHFs. HMGA proteins cooperate with the p16(INK4a) tumor suppressor to promote SAHF formation and proliferative arrest and stabilize senescence by contributing to the repression of proliferation-associated genes. These antiproliferative activities are canceled by coexpression of the HDM2 and CDK4 oncogenes, which are often coamplified with HMGA2 in human cancers. Our results identify a component of the senescence machinery that contributes to heterochromatin formation and imply that HMGA proteins also act in tumor suppressor networks.

Premature proliferative arrest in benign or early-stage tumors induced by oncoproteins, chromosomal instability, or DNA damage is associated with p53/p21 activation, culminating in either senescence or apoptosis, depending on cell context. Growth hormone (GH) elicits direct peripheral metabolic actions as well as growth effects mediated by insulin-like growth factor 1 (IGF1). Locally produced peripheral tissue GH, in contrast to circulating pituitary-derived endocrine GH, has been proposed to be both proapoptotic and prooncogenic. Pituitary adenomas expressing and secreting GH are invariably benign and exhibit DNA damage and a senescent phenotype. We therefore tested effects of nutlin-induced p53-mediated senescence in rat and human pituitary cells. We show that DNA damage senescence induced by nutlin triggers the p53/p21 senescent pathway, with subsequent marked induction of intracellular pituitary GH in vitro. In contrast, GH is not induced in cells devoid of p53. Furthermore we show that p53 binds specific GH promoter motifs and enhances GH transcription and secretion in senescent pituitary adenoma cells and also in nonpituitary (human breast and colon) cells. In vivo, treatment with nutlin results in up-regulation of both p53 and GH in the pituitary gland, as well as increased GH expression in nonpituitary tissues (lung and liver). Intracrine GH acts in pituitary cells as an apoptosis switch for p53-mediated senescence, likely protecting the pituitary adenoma from progression to malignancy. Unlike in the pituitary, in nonpituitary cells GH exerts antiapoptotic properties. Thus, the results show that GH is a direct p53 transcriptional target and fulfills criteria as a p53 target gene. Induced GH is a readily measurable cell marker for p53-mediated cellularsenescence.

Different anatomic and physiological changes occur in the lung of aging people that can affect pulmonary functions, and different pulmonary diseases, including deadly diseases such as chronic obstructive pulmonary disease (COPD)/emphysema and idiopathic pulmonary fibrosis (IPF), can be related to an acceleration of the aging process. The individual genetic background, as well as exposure to a variety of toxic substances (cigarette smoke in primis) can contribute significantly to accelerating pulmonary senescence. Premature aging can impair lung function by different ways: by interfering specifically with tissue repair mechanisms after damage, thus perturbing the correct crosstalk between mesenchymal and epithelial components; by inducing systemic and/or local alteration of the immune system, thus impairing the complex mechanisms of lung defense against infections; and by stimulating a local and/or systemic inflammatory condition (inflammaging). According to recently proposed pathogenic models in COPD and IPF, premature cellularsenescence likely affects distinct progenitors cells (mesenchymal stem cells in COPD, alveolar epithelial precursors in IPF), leading to stem cell exhaustion. In this review, the large amount of data supporting this pathogenic view are discussed, with emphasis on the possible molecular and cellular mechanisms leading to the severe parenchymal remodeling that characterizes, in different ways, these deadly diseases.

Subtelomeres are patchworks of evolutionary conserved sequence blocks and harbor the transcriptional start sites for telomere repeat containing RNAs (TERRA). Recent studies suggest that the interplay between telomeres and subtelomeric chromatin is required for maintaining telomere function. To further characterize chromatin remodeling of subtelomeres in relation to telomere shortening and cellularsenescence, we systematically quantified histone modifications and DNA methylation at the subtelomeres of chromosomes 7q and 11q in primary human WI-38 fibroblasts. Upon senescence, both subtelomeres were characterized by a decrease in markers of constitutive heterochromatin, suggesting relative chromatin relaxation. However, we did not find increased levels of markers of euchromatin or derepression of the 7q VIPR2 gene. The repressed state of the subtelomeres was maintained upon senescence, which could be attributed to a rise in levels of facultative heterochromatin markers at both subtelomeres. While senescence-induced subtelomeric chromatin remodeling was similar for both chromosomes, chromatin remodeling at TERRA promoters displayed chromosome-specific patterns. At the 7q TERRA promoter, chromatin structure was co-regulated with the more proximal subtelomere. In contrast, the 11q TERRA promoter, which was previously shown to be bound by CCCTC-binding factor CTCF, displayed lower levels of markers of constitutive heterochromatin that did not change upon senescence, whereas levels of markers of facultative heterochromatin decreased upon senescence. In line with the chromatin state data, transcription of 11q TERRA but not 7q TERRA was detected. Our study provides a detailed description of human subtelomeric chromatin dynamics and shows distinct regulation of the TERRA promoters of 7q and 11q upon cellularsenescence. PMID:23644601

The process of cellularsenescence generates a repressive chromatin environment, however, the role of histone variants and histone proteolytic cleavage in senescence remains unclear. Using models of oncogene-induced and replicative senescence, here we report novel histone H3 tail cleavage events mediated by the protease Cathepsin L. We find that cleaved forms of H3 are nucleosomal and the histone variant H3.3 is the preferred cleaved form of H3. Ectopic expression of H3.3 and its cleavage product (H3.3cs1), which lacks the first twenty-one amino acids of the H3 tail, is sufficient to induce senescence. Further, H3.3cs1 chromatin incorporation is mediated by the HUCA histone chaperone complex. Genome-wide transcriptional profiling revealed that H3.3cs1 facilitates transcriptional silencing of cell cycle regulators including RB/E2F target genes, likely via the permanent removal of H3K4me3. Collectively, our study identifies histone H3.3 and its proteolytically processed forms as key regulators of cellularsenescence. PMID:25394905

The population of baby boomers (age 60-65) is rapidly increasing globally. The aging of the human body is associated with the decline of cellular function which leads to the development of a variety of diseases. The increased demand for health care for the aging population creates significant financial burden to any healthcare system. Developing strategies and health intervention methods to ameliorate this situation is paramount. Experiments utilizing heterochronic parabiosis in mice have demonstrated that replacing the aging cellular milieu with the plasma of a young experimental animal leads to reversal of cellularsenescence. This article describes a hypothetical model of intermittent heterochronic plasma exchange in humans as a modality for heterochronic parabiosis in an attempt to delay cellularsenescence.

High-affinity sodium-dependent dicarboxylate cotransporter 3 (NaDC3) is a key metabolism-regulating membrane protein responsible for transport of Krebs cycle intermediates. NaDC3 is upregulated as organs age, but knowledge regarding the underlying mechanisms by which NaDC3 modulates mammalian aging is limited. In this study, we showed that NaDC3 overexpression acceleratedcellularsenescence in young human diploid cells (MRC-5 and WI-38) and primary renal tubular cells, leading to cell cycle arrest in G1 phase and increased expression of senescent biomarkers, senescence-associated β-galactosidase and p16. Intracellular levels of reactive oxygen species, 8-hydroxy-2'-deoxyguanosine, malondialdehyde, and carbonyl were significantly enhanced, and activities of respiratory complexes I and III and ATP level were significantly decreased in NaDC3-infected cells. Stressful premature senescent phenotypes induced by NaDC3 were markedly ameliorated via treatment with the antioxidants Tiron and Tempol. High expression of NaDC3 caused a prominent increase in intracellular levels of Krebs cycle intermediates and NADH. Exogenous NADH and NAD(+) may aggravate and attenuate the aging phenotypes induced by NaDC3, respectively. These results suggest that NaDC3 can induce premature cellularsenescence by promoting the transport of Krebs cycle intermediates, increasing generation of NADH and reactive oxygen species and leading to oxidative damage. Our results clarify the aging signaling pathway regulated by NaDC3.

Highlights: Black-Right-Pointing-Pointer Phenylbutyric acid induces cellularsenescence. Black-Right-Pointing-Pointer Phenylbutyric acid activates Akt kinase. Black-Right-Pointing-Pointer The knockdown of PERK also can induce cellularsenescence. Black-Right-Pointing-Pointer Akt/p21{sup WAF1} pathway activates in PERK knockdown induced cellularsenescence. -- Abstract: It has been well known that three sentinel proteins - PERK, ATF6 and IRE1 - initiate the unfolded protein response (UPR) in the presence of misfolded or unfolded proteins in the ER. Recent studies have demonstrated that upregulation of UPR in cancer cells is required to survive and proliferate. Here, we showed that long exposure to 4-phenylbutyric acid (PBA), a chemical chaperone that can reduce retention of unfolded and misfolded proteins in ER, induced cellularsenescence in cancer cells such as MCF7 and HT1080. In addition, we found that treatment with PBA activates Akt, which results in p21{sup WAF1} induction. Interestingly, the depletion of PERK but not ATF6 and IRE1 also induces cellularsenescence, which was rescued by additional depletion of Akt. This suggests that Akt pathway is downstream of PERK in PBA induced cellularsenescence. Taken together, these results show that PBA induces cellularsenescence via activation of the Akt/p21{sup WAF1} pathway by PERK inhibition.

Cellularsenescence arrests the proliferation of mammalian cells at risk for neoplastic transformation, and is also associated with aging. However, the factors that cause cellularsenescence during aging are unclear. Excessive reactive oxygen species (ROS) have been shown to cause cellularsenescence in culture, and accumulated molecular damage due to mitochondrial ROS has long been thought to drive aging phenotypesin vivo. Here, we test the hypothesis that mitochondrial oxidative stress can promote cellularsenescence in vivo and contribute to aging phenotypes in vivo, specifically in the skin. We show that the number of senescent cells, as well as impaired mitochondrial (complex II) activity increase in naturally aged mouse skin. Using a mouse model of genetic Sod2 deficiency, we show that failure to express this important mitochondrial anti-oxidant enzyme also impairs mitochondrial complex II activity, causes nuclear DNA damage, and induces cellularsenescence but not apoptosis in the epidermis. Sod2 deficiency also reduced the number of cells and thickness of the epidermis, while increasing terminal differentiation. Our results support the idea that mitochondrial oxidative stress and cellularsenescence contribute to aging skin phenotypes in vivo.

Cells exposed to genotoxic stress induce cellularsenescence through a DNA damage response (DDR) pathway regulated by ATM kinase and reactive oxygen species (ROS). Here, we show that the regulatory roles for ATM kinase and ROS differ during induction and maintenance of cellularsenescence. Cells treated with different genotoxic agents were analyzed using specific pathway markers and inhibitors to determine that ATM kinase activation is directly proportional to the dose of the genotoxic stress and that senescence initiation is not dependent on ROS or the p53 status of cells. Cells in which ROS was quenched still activated ATM and initiated the DDR when insulted, and progressed normally to senescence. By contrast, maintenance of a viable senescent state required the presence of ROS as well as activated ATM. Inhibition or removal of either of the components caused cell death in senescent cells, through a deregulated ATM-ROS axis. Overall, our work demonstrates existence of an intricate temporal hierarchy between genotoxic stress, DDR and ROS in cellularsenescence. Our model reports the existence of different stages of cellularsenescence with distinct regulatory networks.

Hutchinson-Gilford Progeria Syndrome (HGPS) is a premature-aging syndrome caused by a dominant mutation in the gene encoding lamin A, which leads to an aberrantly spliced and processed protein termed progerin. Previous studies have shown that progerin induces early senescence associated with increased DNA-damage signaling and that telomerase extends HGPS cellular lifespan. We demonstrate that telomerase extends HGPS cellular lifespan by decreasing progerin-induced DNA-damage signaling and activation of p53 and Rb pathways that otherwise mediate the onset of premature senescence. We show further that progerin-induced DNA-damage signaling is localized to telomeres and is associated with telomere aggregates and chromosomal aberrations. Telomerase amelioration of DNA-damage signaling is relatively rapid, requires both its catalytic and DNA-binding functions, and correlates in time with the acquisition by HGPS cells of the ability to proliferate. All of these findings establish that HGPS premature cellularsenescence results from progerin-induced telomere dysfunction.

Cellularsenescence is an irreversible cell cycle arrest in which specific mRNAs and miRNAs are involved in senescence progression. miRNAs interact with specific mRNAs to regulate various cellular mechanisms, including metabolism, proliferation, apoptosis, senescence and differentiation. In this study, we identify and characterize miRNAs during cellularsenescence in mesenchymal stem cells (MSCs). Using previously reported miRNAs, expression profiling of 23 miRNAs was performed using real-time PCR analysis. Among these miRNAs, 19 miRNAs showed upregulated expression patterns in senescent MSCs compared with young MSCs, and 5 miRNAs were downregulated. These miRNAs have not been previously identified as being related to cellularsenescence but seem to be related. miR-103-2*, miR-140-5p and miR-330-5p are highly upregulated, while miR-29b and miR-199b-5p are significantly downregulated in senescent MSCs. We identify unique functions of 5 miRNAs and predict putative target genes of 5 miRNAs using our previous report. Among them, miR-199b-5p directly suppressed LAMC1 expression, as shown in a luciferase assay. miR-199b-5p significantly regulates translational activity but does not control post-transcriptional activity. Likewise, miR-199b-5p modulates LAMC networks, which demonstrates the resulting phenomenon during cellularsenescence, namely, that miR-199b-5p indirectly regulates cellularsenescence in MSCs.

The KPs (kisspeptins) are a family of multifunctional peptides with established roles in cancer metastasis, puberty and vasoconstriction. The effects of KPs on endothelial cells have yet to be determined. The aim of the present study was to investigate the effects of KP-10 on endothelial cell growth and the mechanisms underlying those effects. The administration of recombinant KP-10 into the hindlimbs of rats with ischaemia significantly impaired blood flow recovery, as shown by laser Doppler, and capillary growth, as shown using histology, compared with the controls. HUVECs (human umbilical vein endothelial cells) express the KP receptor and were treated with KP-10 in culture studies. KP-10 inhibited endothelial cell tube formation and proliferation in a significant and dose-dependent manner. The HUVECs treated with KP exhibited the senescent phenotype, as determined using a senescence-associated β-galactosidase assay, cell morphology analysis, and decreased Sirt1 (sirtuin 1) expression and increased p53 expression shown by Western blot analysis. Intriguingly, a pharmacological Rho kinase inhibitor, Y-27632, was found to increase the proliferation of HUVECs and to reduce the number of senescent phenotype cells affected by KP-10. In conclusion, KP-10 suppressed endothelial cells growth both in vivo and in vitro in the present study. The adverse effect of KP on endothelial cells was attributable, at least in part, to the induction of cellularsenescence.

The cellular and subcellular localization of endogenous nitric oxide (NO.) in leaves from young and senescent pea (Pisum sativum) plants was studied. Confocal laser scanning microscopy analysis of pea leaf sections with the fluorescent probe 4,5-diaminofluorescein diacetate revealed that endogenous NO. was mainly present in vascular tissues (xylem and phloem). Green fluorescence spots were also detected in the epidermal cells, palisade and spongy mesophyll cells, and guard cells. In senescent leaves, NO. generation was clearly reduced in the vascular tissues. At the subcellular level, by electron paramagnetic resonance spectroscopy with the spin trap Fe(MGD)(2) and fluorometric analysis with 4,5-diaminofluorescein diacetate, NO. was found to be an endogenous metabolite of peroxisomes. The characteristic three-line electron paramagnetic resonance spectrum of NO., with g = 2.05 and a(N) = 12.8 G, was detected in peroxisomes. By fluorometry, NO. was also found in these organelles, and the level measured of NO. was linearly dependent on the amount of peroxisomal protein. The enzymatic production of NO. from l-Arg (nitric oxide synthase [NOS]-like activity) was measured by ozone chemiluminiscence. The specific activity of peroxisomal NOS was 4.9 nmol NO. mg(-1) protein min(-1); was strictly dependent on NADPH, calmodulin, and BH(4); and required calcium. In senescent pea leaves, the NOS-like activity of peroxisomes was down-regulated by 72%. It is proposed that peroxisomal NO. could be involved in the process of senescence of pea leaves.

Telomeres are nucleoprotein structures located at the linear ends of eukaryotic chromosomes. Telomere integrity is required for cell proliferation and survival. Although the vast majority of eukaryotic species use telomerase as a primary means for telomere maintenance, a few species can use recombination or retrotransposon-mediated maintenance pathways. Since Saccharomyces cerevisiae can use both telomerase and recombination to replicate telomeres, budding yeast provides a useful system with which to examine the evolutionary advantages of telomerase and recombination in preserving an organism or cell under natural selection. In this study, we examined the life span in telomerase-null, post-senescent type II survivors that have employed homologous recombination to replicate their telomeres. Type II recombination survivors stably maintained chromosomal integrity but exhibited a significantly reduced replicative life span. Normal patterns of cell morphology at the end of a replicative life span and aging-dependent sterility were observed in telomerase-null type II survivors, suggesting the type II survivors aged prematurely in a manner that is phenotypically consistent with that of wild-type senescent cells. The shortened life span of type II survivors was extended by calorie restriction or TOR1 deletion, but not by Fob1p inactivation or Sir2p over-expression. Intriguingly, rDNA recombination was decreased in type II survivors, indicating that the premature aging of type II survivors was not caused by an increase in extra-chromosomal rDNA circle accumulation. Reintroduction of telomerase activity immediately restored the replicative life span of type II survivors despite their heterogeneous telomeres. These results suggest that telomere recombination acceleratescellular aging in telomerase-null type II survivors and that telomerase is likely a superior telomere maintenance pathway in sustaining yeast replicative life span. PMID:19557187

Human respiratory syncytial virus (HRSV) accounts for the majority of lower respiratory tract infections during infancy and childhood and is associated with significant morbidity and mortality. HRSV provokes a proliferation arrest and characteristic syncytia in cellular systems such as immortalized epithelial cells. We show here that HRSV induces the expression of DNA damage markers and proliferation arrest such as P-TP53, P-ATM, CDKN1A and γH2AFX in cultured cells secondary to the production of mitochondrial reactive oxygen species (ROS). The DNA damage foci contained γH2AFX and TP53BP1, indicative of double-strand breaks (DSBs) and could be reversed by antioxidant treatments such as N-Acetylcysteine (NAC) or reduced glutathione ethyl ester (GSHee). The damage observed is associated with the accumulation of senescent cells, displaying a canonical senescent phenotype in both mononuclear cells and syncytia. In addition, we show signs of DNA damage and aging such as γH2AFX and CDKN2A expression in the respiratory epithelia of infected mice long after viral clearance. Altogether, these results show that HRSV triggers a DNA damage-mediated cellularsenescence program probably mediated by oxidative stress. The results also suggest that this program might contribute to the physiopathology of the infection, tissue remodeling and aging, and might be associated to long-term consequences of HRSV infections.

ABSTRACT Human respiratory syncytial virus (HRSV) accounts for the majority of lower respiratory tract infections during infancy and childhood and is associated with significant morbidity and mortality. HRSV provokes a proliferation arrest and characteristic syncytia in cellular systems such as immortalized epithelial cells. We show here that HRSV induces the expression of DNA damage markers and proliferation arrest such as P-TP53, P-ATM, CDKN1A and γH2AFX in cultured cells secondary to the production of mitochondrial reactive oxygen species (ROS). The DNA damage foci contained γH2AFX and TP53BP1, indicative of double-strand breaks (DSBs) and could be reversed by antioxidant treatments such as N-Acetylcysteine (NAC) or reduced glutathione ethyl ester (GSHee). The damage observed is associated with the accumulation of senescent cells, displaying a canonical senescent phenotype in both mononuclear cells and syncytia. In addition, we show signs of DNA damage and aging such as γH2AFX and CDKN2A expression in the respiratory epithelia of infected mice long after viral clearance. Altogether, these results show that HRSV triggers a DNA damage-mediated cellularsenescence program probably mediated by oxidative stress. The results also suggest that this program might contribute to the physiopathology of the infection, tissue remodeling and aging, and might be associated to long-term consequences of HRSV infections. PMID:26809688

The coordination of cell cycle progression with the repair of DNA damage supports the genomic integrity of dividing cells. The function of many factors involved in DNA damage response (DDR) and the cell cycle depends on their Ran GTPase–regulated nuclear–cytoplasmic transport (NCT). The loading of Ran with GTP, which is mediated by RCC1, the guanine nucleotide exchange factor for Ran, is critical for NCT activity. However, the role of RCC1 or Ran⋅GTP in promoting cell proliferation or DDR is not clear. We show that RCC1 overexpression in normal cells increased cellular Ran⋅GTP levels and accelerated the cell cycle and DNA damage repair. As a result, normal cells overexpressing RCC1 evaded DNA damage–induced cell cycle arrest and senescence, mimicking colorectal carcinoma cells with high endogenous RCC1 levels. The RCC1-induced inhibition of senescence required Ran and exportin 1 and involved the activation of importin β–dependent nuclear import of 53BP1, a large NCT cargo. Our results indicate that changes in the activity of the Ran⋅GTP–regulated NCT modulate the rate of the cell cycle and the efficiency of DNA repair. Through the essential role of RCC1 in regulation of cellular Ran⋅GTP levels and NCT, RCC1 expression enables the proliferation of cells that sustain DNA damage. PMID:26864624

Oxidative stress and persistent DNA damage response contribute to cellularsenescence, a degeneration process critically involving ataxia telangiectasia-mutated (ATM) and p53. Selenoprotein H (SelH), a nuclear selenoprotein, is proposed to carry redox and transactivation domains. To determine the role of SelH in genome maintenance, shRNA knockdown was employed in human normal and immortalized cell lines. SelH shRNA MRC-5 diploid fibroblasts under ambient O2 displayed a distinct profile of senescence including β-galactosidase expression, autofluorescence, growth inhibition, and ATM pathway activation. Such senescence phenotypes were alleviated in the presence of ATM kinase inhibitors, by p53 shRNA knockdown, or by maintaining the cells under 3% O2. During the course of 5-day recovery, the induction of phospho-ATM on Ser-1981 and γH2AX by H2O2 treatment (20 μm) subsided in scrambled shRNA but exacerbated in SelH shRNA MRC-5 cells. Results from clonogenic assays demonstrated hypersensitivity of SelH shRNA HeLa cells to paraquat and H2O2, but not to hydroxyurea, neocarzinostatin, or camptothecin. While SelH mRNA expression was induced by H2O2 treatment, SelH-GFP did not mobilize to sites of oxidative DNA damage. The glutathione level was lower in SelH shRNA than scrambled shRNA HeLa cells, and the H2O2-induced cell death was rescued in the presence of N-acetylcysteine, a glutathione precursor. Altogether, SelH protects against cellularsenescence to oxidative stress through a genome maintenance pathway involving ATM and p53. PMID:25336634

Tissue homeostasis requires a carefully-orchestrated balance between cell proliferation, cellularsenescence and cell death. Cells proliferate through a cell cycle that is tightly regulated by cyclin-dependent kinase activities. Cellularsenescence is a safeguard program limiting the proliferative competence of cells in living organisms. Apoptosis eliminates unwanted cells by the coordinated activity of gene products that regulate and effect cell death. The intimate link between the cell cycle, cellularsenescence, apoptosis regulation, cancer development and tumor responses to cancer treatment has become eminently apparent. Extensive research on tumor suppressor genes, oncogenes, the cell cycle and apoptosis regulatory genes has revealed how the DNA damage-sensing and -signaling pathways, referred to as the DNA-damage response network, are tied to cell proliferation, cell-cycle arrest, cellularsenescence and apoptosis. DNA-damage responses are complex, involving "sensor" proteins that sense the damage, and transmit signals to "transducer" proteins, which, in turn, convey the signals to numerous "effector" proteins implicated in specific cellular pathways, including DNA repair mechanisms, cell-cycle checkpoints, cellularsenescence and apoptosis. The Bcl-2 family of proteins stands among the most crucial regulators of apoptosis and performs vital functions in deciding whether a cell will live or die after cancer chemotherapy and irradiation. In addition, several studies have now revealed that members of the Bcl-2 family also interface with the cell cycle, DNA repair/recombination and cellularsenescence, effects that are generally distinct from their function in apoptosis. In this review, we report progress in understanding the molecular networks that regulate cell-cycle checkpoints, cellularsenescence and apoptosis after DNA damage, and discuss the influence of some Bcl-2 family members on cell-cycle checkpoint regulation.

By activating the ataxia telangiectasia mutated (ATM)-mediated DNA Damage Response (DDR), the AMPK agonist metformin might sensitize cells against further damage, thus mimicking the precancerous stimulus that induces an intrinsic barrier against carcinogenesis. Herein, we present the new hypothesis that metformin might function as a tissue sweeper of pre-malignant cells before they gain stem cell/tumor initiating properties. Because enhanced glycolysis (the Warburg effect) plays a causal role in the gain of stem-like properties of tumor-initiating cells by protecting them from the pro-senescent effects of mitochondrial respiration-induced oxidative stress, metformin's ability to disrupt the glycolytic metabotype may generate a cellular phenotype that is metabolically protected against immortalization. The bioenergetic crisis imposed by metformin, which may involve enhanced mitochondrial biogenesis and oxidative stress, can lower the threshold for cellularsenescence by pre-activating an ATM-dependent pseudo-DDR. This allows an accelerated onset of cellularsenescence in response to additional oncogenic stresses. By pushing cancer cells to use oxidative phosphorylation instead of glycolysis, metformin can rescue cell surface major histocompatibility complex class I (MHC-I) expression that is downregulated by oncogenic transformation, a crucial adaptation of tumor cells to avoid the adaptive immune response by cytotoxic T-lymphocytes (CTLs). Aside from restoration of tumor immunosurveillance at the cell-autonomous level, metformin can activate a senescence-associated secretory phenotype (SASP) to reinforce senescence growth arrest, which might trigger an immune-mediated clearance of the senescent cells in a non-cell-autonomous manner. By diminishing the probability of escape from the senescence anti-tumor barrier, the net effect of metformin should be a significant decrease in the accumulation of dysfunctional, pre-malignant cells in tissues, including those with the

Cellularsenescence suppresses cancer by preventing the proliferation of cells that experience potentially oncogenic stimuli. Senescent cells often express p16INK4a, a cyclin-dependent kinase inhibitor, tumor suppressor, and biomarker of aging, which renders the senescence growth arrest irreversible. Senescent cells also acquire a complex phenotype that includes the secretion of many cytokines, growth factors, and proteases, termed a senescence-associated secretory phenotype (SASP). The SASP is proposed to underlie age-related pathologies, including, ironically, late life cancer. Here, we show that ectopic expression of p16INK4a and another cyclin-dependent kinase inhibitor, p21CIP1/WAF1, induces senescence without a SASP, even though they induced other features of senescence, including a stable growth arrest. Additionally, human fibroblasts induced to senesce by ionizing radiation or oncogenic RAS developed a SASP regardless of whether they expressed p16INK4a. Cells induced to senesce by ectopic p16INK4a expression lacked paracrine activity on epithelial cells, consistent with the absence of a functional SASP. Nonetheless, expression of p16INK4a by cells undergoing replicative senescence limited the accumulation of DNA damage and premature cytokine secretion, suggesting an indirect role for p16INK4a in suppressing the SASP. These findings suggest that p16INK4a-positive cells may not always harbor a SASP in vivo and, furthermore, that the SASP is not a consequence of p16INK4a activation or senescence per se, but rather is a damage response that is separable from the growth arrest. PMID:21880712

The plant G-protein network, comprising Gα, Gβ, and Gγ core subunits, regulates development, senses sugar, and mediates biotic and abiotic stress responses. Here, we report G-protein signalling in the salt stress response using two crop models, rice and maize. Loss-of-function mutations in the corresponding genes encoding the Gα subunit attenuate growth inhibition and cellularsenescence caused by sodium chloride (NaCl). Gα null mutations conferred reduced leaf senescence, chlorophyll degradation, and cytoplasm electrolyte leakage under NaCl stress. Sodium accumulated in both wild-type and Gα-mutant shoots to the same levels, suggesting that Gα signalling controls cell death in leaves rather than sodium exclusion in roots. Growth inhibition is probably initiated by osmotic change around root cells, because KCl and MgSO4 also suppressed seedling growth equally as well as NaCl. NaCl lowered rates of cell division and elongation in the wild-type leaf sheath to the level of the Gα-null mutants; however there was no NaCl-induced decrease in cell division in the Gα mutant, implying that the osmotic phase of salt stress suppresses cell proliferation through the inhibition of Gα-coupled signalling. These results reveal two distinct functions of Gα in NaCl stress in these grasses: attenuation of leaf senescence caused by sodium toxicity in leaves, and cell cycle regulation by osmotic/ionic stress.

The plant G-protein network, comprising Gα, Gβ, and Gγ core subunits, regulates development, senses sugar, and mediates biotic and abiotic stress responses. Here in this paper, we report G-protein signalling in the salt stress response using two crop models, rice and maize. Loss-of-function mutations in the corresponding genes encoding the Gα subunit attenuate growth inhibition and cellularsenescence caused by sodium chloride (NaCl). Gα null mutations conferred reduced leaf senescence, chlorophyll degradation, and cytoplasm electrolyte leakage under NaCl stress. Sodium accumulated in both wild-type and Gα-mutant shoots to the same levels, suggesting that Gα signalling controls cell death in leavesmore » rather than sodium exclusion in roots. Growth inhibition is probably initiated by osmotic change around root cells, because KCl and MgSO4 also suppressed seedling growth equally as well as NaCl. NaCl lowered rates of cell division and elongation in the wild-type leaf sheath to the level of the Gα-null mutants; however there was no NaCl-induced decrease in cell division in the Gα mutant, implying that the osmotic phase of salt stress suppresses cell proliferation through the inhibition of Gα-coupled signalling. These results reveal two distinct functions of Gα in NaCl stress in these grasses: attenuation of leaf senescence caused by sodium toxicity in leaves, and cell cycle regulation by osmotic/ionic stress.« less

The plant G-protein network, comprising Gα, Gβ, and Gγ core subunits, regulates development, senses sugar, and mediates biotic and abiotic stress responses. Here in this paper, we report G-protein signalling in the salt stress response using two crop models, rice and maize. Loss-of-function mutations in the corresponding genes encoding the Gα subunit attenuate growth inhibition and cellularsenescence caused by sodium chloride (NaCl). Gα null mutations conferred reduced leaf senescence, chlorophyll degradation, and cytoplasm electrolyte leakage under NaCl stress. Sodium accumulated in both wild-type and Gα-mutant shoots to the same levels, suggesting that Gα signalling controls cell death in leaves rather than sodium exclusion in roots. Growth inhibition is probably initiated by osmotic change around root cells, because KCl and MgSO4 also suppressed seedling growth equally as well as NaCl. NaCl lowered rates of cell division and elongation in the wild-type leaf sheath to the level of the Gα-null mutants; however there was no NaCl-induced decrease in cell division in the Gα mutant, implying that the osmotic phase of salt stress suppresses cell proliferation through the inhibition of Gα-coupled signalling. These results reveal two distinct functions of Gα in NaCl stress in these grasses: attenuation of leaf senescence caused by sodium toxicity in leaves, and cell cycle regulation by osmotic/ionic stress.

The plant G-protein network, comprising Gα, Gβ, and Gγ core subunits, regulates development, senses sugar, and mediates biotic and abiotic stress responses. Here, we report G-protein signalling in the salt stress response using two crop models, rice and maize. Loss-of-function mutations in the corresponding genes encoding the Gα subunit attenuate growth inhibition and cellularsenescence caused by sodium chloride (NaCl). Gα null mutations conferred reduced leaf senescence, chlorophyll degradation, and cytoplasm electrolyte leakage under NaCl stress. Sodium accumulated in both wild-type and Gα-mutant shoots to the same levels, suggesting that Gα signalling controls cell death in leaves rather than sodium exclusion in roots. Growth inhibition is probably initiated by osmotic change around root cells, because KCl and MgSO4 also suppressed seedling growth equally as well as NaCl. NaCl lowered rates of cell division and elongation in the wild-type leaf sheath to the level of the Gα-null mutants; however there was no NaCl-induced decrease in cell division in the Gα mutant, implying that the osmotic phase of salt stress suppresses cell proliferation through the inhibition of Gα-coupled signalling. These results reveal two distinct functions of Gα in NaCl stress in these grasses: attenuation of leaf senescence caused by sodium toxicity in leaves, and cell cycle regulation by osmotic/ionic stress. PMID:25227951

Research highlights: {yields} Resveratrol induces cellularsenescence in glioma cell. {yields} Resveratrol inhibits mono-ubiquitination of histone H2B at K120. {yields} Depletion of RNF20, phenocopies the inhibitory effects of resveratrol. {yields} Mono-ubiquitination of histone H2B at K120 is a novel target of resveratrol. {yields} RNF20 inhibits cellularsenescence in proliferating glioma cells. -- Abstract: Resveratrol (3,4',5-trihydroxy-trans-stilbene), a polyphenol naturally occurring in grapes and other plants, has cancer chemo-preventive effects and therapeutic potential. Although resveratrol modulates multiple pathways in tumor cells, how resveratrol or its affected pathways converge on chromatin to mediate its effects is not known. Using glioma cells as a model, we showed here that resveratrol inhibited cell proliferation and induced cellular hypertrophy by transforming spindle-shaped cells to enlarged, irregular and flatten-shaped ones. We further showed that resveratrol-induced hypertrophic cells expressed senescence-associated-{beta}-galactosidase, suggesting that resveratrol-induced cellularsenescence in glioma cells. Consistent with these observations, we demonstrated that resveratrol inhibited clonogenic efficiencies in vitro and tumor growth in a xenograft model. Furthermore, we found that acute treatment of resveratrol inhibited mono-ubiquitination of histone H2B at K120 (uH2B) in breast, prostate, pancreatic, lung, brain tumor cells as well as primary human cells. Chronic treatment with low doses of resveratrol also inhibited uH2B in the resveratrol-induced senescent glioma cells. Moreover, we showed that depletion of RNF20, a ubiquitin ligase of histone H2B, inhibited uH2B and induced cellularsenescence in glioma cells in vitro, thereby recapitulated the effects of resveratrol. Taken together, our results suggest that uH2B is a novel direct or indirect chromatin target of resveratrol and RNF20 plays an important role in inhibiting cellular

Highlights: • Dyskerin depletion triggers cellularsenescence in U2OS osteosarcoma cells. • Dyskerin-depleted cells are resistant to apoptosis induced by genotoxic stress. • Chromatin relaxation sensitizes dyskerin-depleted cells to apoptosis. - Abstract: Dyskerin is a conserved, nucleolar RNA-binding protein implicated in an increasing array of fundamental cellular processes. Germline mutation in the dyskerin gene (DKC1) is the cause of X-linked dyskeratosis congenita (DC). Conversely, wild-type dyskerin is overexpressed in sporadic cancers, and high-levels may be associated with poor prognosis. It was previously reported that acute loss of dyskerin function via siRNA-mediated depletion slowed the proliferation of transformed cell lines. However, the mechanisms remained unclear. Using human U2OS osteosarcoma cells, we show that siRNA-mediated dyskerin depletion induced cellularsenescence as evidenced by proliferative arrest, senescence-associated heterochromatinization and a senescence-associated molecular profile. Senescence can render cells resistant to apoptosis. Conversely, chromatin relaxation can reverse the repressive effects of senescence-associated heterochromatinization on apoptosis. To this end, genotoxic stress-induced apoptosis was suppressed in dyskerin-depleted cells. In contrast, agents that induce chromatin relaxation, including histone deacetylase inhibitors and the DNA intercalator chloroquine, sensitized dyskerin-depleted cells to apoptosis. Dyskerin is a core component of the telomerase complex and plays an important role in telomere homeostasis. Defective telomere maintenance resulting in premature senescence is thought to primarily underlie the pathogenesis of X-linked DC. Since U2OS cells are telomerase-negative, this leads us to conclude that loss of dyskerin function can also induce cellularsenescence via mechanisms independent of telomere shortening.

Acute kidney injury is a major clinical problem and advanced age is associated with ineffective renal regeneration and poor functional outcome. Data from kidney injury models suggest that a loss of tubular epithelial proliferation contributes to a decrease in renal repair capacity with aging, but aging can also lead to a higher severity of inflammation and damage which may influence repair. In this study we tested intrinsic age-dependent changes in tubular epithelial proliferation in young and old mice, by injecting low-dose lead acetate as a non-injurious mitogen. In parallel, we explored in vitro techniques of studying cellularsenescence in primary tubular epithelial cells (PTEC). Lead acetate induced tubular epithelial proliferation at a significantly higher rate in young as compared to old mice. Old kidneys showed significantly more senescence as demonstrated by increased p16 (INK4a), senescence associated β-galactosidase, and γH2AX(+)/Ki-67(-) cells. This was paralleled in old kidneys by a higher number of Cyclin D1 positive tubular cells. This finding was corroborated by a positive correlation between Cyclin D1 positivity and age in human renal biopsies. When tubular cells were isolated from mouse kidneys they rapidly lost their age-associated differences under culture conditions. However, senescence was readily induced in PTEC by γ-irradiation representing a future model for study of cellularsenescence in the renal epithelium. Together, our data indicate that the tubular epithelium of aged kidney has an intrinsically reduced proliferative capacity probably due to a higher load of senescent cells. Moreover, stress induced models of cellularsenescence are preferable for study of the renal epithelium in vitro. Finally, the positive correlation of Cyclin D1 with age and cellularsenescence in PTEC needs further evaluation as to a functional role of renal epithelial aging.

Background Micro RNAs (miRs) have emerged as key regulators during oncogenesis. They have been found to regulate cell proliferation, differentiation, and apoptosis. Mir-125b has been identified as an oncomir in various forms of tumours, but we have previously proposed that miR-125b is a suppressor of lymph node metastasis in cutaneous malignant melanoma. Our goal was therefore to further examine this theory. Methods We used in-situ-hybridization to visualise miR-125b expression in primary tumours and in lymph node metastasis. Then using a miRVector plasmid containing a miR-125b-1 insert we transfected melanoma cell line Mel-Juso and then investigated the effect of the presence of a stable overexpression of miR-125b on growth by western blotting, flow cytometry and β-galactosidase staining. The tumourogenicity of the transfected cells was tested using a murine model and the tumours were further examined with in-situ-hybridization. Results In primary human tumours and in lymph node metastases increased expression of miR-125b was found in single, large tumour cells with abundant cytoplasm. A stable overexpression of miR-125b in human melanoma cell line Mel-Juso resulted in a G0/G1 cell cycle block and emergence of large cells expressing senescence markers: senescence-associated beta-galactosidase, p21, p27 and p53. Mel-Juso cells overexpressing miR-125b were tumourigenic in mice, but the tumours exhibited higher level of cell senescence and decreased expression of proliferation markers, cyclin D1 and Ki67 than the control tumours. Conclusions Our results confirm the theory that miR-125b functions as a tumour supressor in cutaneous malignant melanoma by regulating cellularsenescence, which is one of the central mechanisms protecting against the development and progression of malignant melanoma. PMID:24762088

Cellularsenescence is an irreversible cell cycle arrest that limits the replicative lifespan of cells. Senescence suppresses development of tumors by regulating aging factors, such as cyclin dependent kinase inhibitor (CKI) and telomerase. Suppression subtractive hybridization (SSH) was used to identify genes that were differentially expressed between young human mesenchymal stem cells (Y-hMSCs) and senescent human mesenchymal stem cells (S-hMSCs). We selected positive clones that were functionally characterized by referring to public databases using NCBI BLAST tool. This search revealed that 19 genes were downregulated, and 43 genes were upregulated in S-hMSCs relative to Y-hMSCs. Among subtracted clones in Y-hMSCs, most of genes markedly were related to metabolic functions. These genes, PDIA3, WDR1, FSTL1, COPG1, LMAN1, and PDIA6, significantly downregulated. Conversely, genes for subtracted clones in S-hMSCs were mostly associated with cell adhesion. In particular, the expression levels of 9 genes, HSP90B1, EID1, ATP2B4, DDAH1, PRNP, RAB1A, PGS5, TM4SF1 and SSR3, gradually increased during senescence. These genes have not previously been identified as being related to cellularsenescence, but they seemed to be potentially affected during cellularsenescence.

LMNA mutations are responsible for a variety of genetic disorders, including muscular dystrophy, lipodystrophy, and certain progeroid syndromes, notably Hutchinson-Gilford Progeria. Although a number of clinical features of these disorders are suggestive of accelerated aging, it is not known whether cells derived from these patients exhibit cellular phenotypes associated with accelerated aging. We examined a series of isogenic skin fibroblast lines transfected with LMNA constructs bearing known pathogenic point mutations or deletion mutations found in progeroid syndromes. Fibroblasts overexpressing mutant lamin A exhibited accelerated rates of loss of telomeres and shortened replicative lifespans, in addition to abnormal nuclear morphology. To our surprise, these abnormalities were also observed in lines overexpressing wild-type lamin A. Copy number variants are common in human populations; those involving LMNA, whether arising meiotically or mitotically, might lead to progeroid phenotypes. In an initial pilot study of 23 progeroid cases without detectable WRN or LMNA mutations, however, no cases of altered LMNA copy number were detected. Nevertheless, our findings raise a hypothesis that changes in lamina organization may cause accelerated telomere attrition, with different kinetics for overexpession of wild-type and mutant lamin A, which leads to rapid replicative senescence and progroid phenotypes.

Ultraviolet (UV) irradiation causes damage in skin by generating excessive reactive oxygen species (ROS) and induction of matrix metalloproteinases (MMPs), leading to skin photoageing. Dandelion extracts have long been used for traditional Chinese medicine and native American medicine to treat cancers, hepatitis, and digestive diseases; however, less is known on the effects of dandelion extracts in skin photoageing. Here we found that dandelion leaf and flower extracts significantly protect UVB irradiation-inhibited cell viability when added before UVB irradiation or promptly after irradiation. Dandelion leaf and flower extracts inhibited UVB irradiation-stimulated MMP activity and ROS generation. Dandelion root extracts showed less action on protecting HDFs from UVB irradiation-induced MMP activity, ROS generation, and cell death. Furthermore, dandelion leaf and flower but not root extracts stimulated glutathione generation and glutathione reductase mRNA expression in the presence or absence of UVB irradiation. We also found that dandelion leaf and flower extracts help absorb UVB irradiation. In addition, dandelion extracts significantly protected HDFs from H2O2-induced cellularsenescence. In conclusion, dandelion extracts especially leaf and flower extracts are potent protective agents against UVB damage and H2O2-induced cellularsenescence in HDFs by suppressing ROS generation and MMP activities and helping UVB absorption. PMID:26576225

The role of Sirtuin 6 (SIRT6) as a tumor suppressor or oncogene in liver cancer remains controversial. Thus, we identified the specific role of SIRT6 in the progression of hepatocellular carcinoma (HCC). SIRT6 expression was significantly higher in HCC cell lines and HCC tissues from 138 patients than in an immortalized hepatocyte cell line, THLE-2 and non-tumor tissues, respectively. SIRT6 knockdown by shRNA suppressed the growth of HCC cells and inhibited HCC tumor growth in vivo. In addition, SIRT6 silencing significantly prevented the growth of HCC cell lines by inducing cellularsenescence in the p16/Rb- and p53/p21-pathway independent manners. Microarray analysis revealed that the expression of genes involved in nucleosome assembly was apparently altered in SIRT6-depleted Hep3B cells. SIRT6 knockdown promoted G2/M phase arrest and downregulation of genes encoding histone variants associated with nucleosome assembly, which could be attributed to DNA damage. Taken together, our findings suggest that SIRT6 acts as a tumor promoter by preventing DNA damage and cellularsenescence, indicating that SIRT6 represents a potential therapeutic target for the treatment of HCC. PMID:27824900

Ultraviolet (UV) irradiation causes damage in skin by generating excessive reactive oxygen species (ROS) and induction of matrix metalloproteinases (MMPs), leading to skin photoageing. Dandelion extracts have long been used for traditional Chinese medicine and native American medicine to treat cancers, hepatitis, and digestive diseases; however, less is known on the effects of dandelion extracts in skin photoageing. Here we found that dandelion leaf and flower extracts significantly protect UVB irradiation-inhibited cell viability when added before UVB irradiation or promptly after irradiation. Dandelion leaf and flower extracts inhibited UVB irradiation-stimulated MMP activity and ROS generation. Dandelion root extracts showed less action on protecting HDFs from UVB irradiation-induced MMP activity, ROS generation, and cell death. Furthermore, dandelion leaf and flower but not root extracts stimulated glutathione generation and glutathione reductase mRNA expression in the presence or absence of UVB irradiation. We also found that dandelion leaf and flower extracts help absorb UVB irradiation. In addition, dandelion extracts significantly protected HDFs from H2O2-induced cellularsenescence. In conclusion, dandelion extracts especially leaf and flower extracts are potent protective agents against UVB damage and H2O2-induced cellularsenescence in HDFs by suppressing ROS generation and MMP activities and helping UVB absorption.

The amount of cellular proteins is a crucial parameter that is known to vary between cells as a function of the replicative passages, and can be important during physiological aging. The process of protein degradation is known to be performed by a series of enzymatic reactions, ranging from an initial step of protein ubiquitination to their final fragmentation by the proteasome. In this paper we propose a stochastic dynamical model of nuclear proteins concentration resulting from a balance between a constant production of proteins and their degradation by a cooperative enzymatic reaction. The predictions of this model are compared with experimental data obtained by fluorescence measurements of the amount of nuclear proteins in murine tail fibroblast (MTF) undergoing cellularsenescence. Our model provides a three-parameter stationary distribution that is in good agreement with the experimental data even during the transition to the senescent state, where the nuclear protein concentration changes abruptly. The estimation of three parameters (cooperativity, saturation threshold, and maximal velocity of the reaction), and their evolution during replicative passages shows that only the maximal velocity varies significantly. Based on our modeling we speculate the reduction of functionality of the protein degradation mechanism as a possible competitive inhibition of the proteasome. PMID:26115222

S-phase kinase-associated protein 2 (Skp2) belongs to the F-box protein family. It is a component of the SCF E3 ubiquitin ligase complex. Skp2 has been shown to regulate cellular proliferation by targeting several cell cycle-regulated proteins for ubiquitination and degradation, including cyclin-dependent kinase inhibitor p27. Skp2 has also been demonstrated to display an oncogenic function since its overexpression has been observed in many human cancers. This review discusses the recent discoveries on the novel roles of Skp2 in regulating cellularsenescence, cancer progression, and metastasis, as well as the therapeutic potential of targeting Skp2 for human cancer treatment. PMID:22200179

Osteoporosis and avascular necrosis (AVN) are long-lasting and debilitating complications of hematopoietic stem cell transplantation (HSCT). We describe the magnitude of bone loss, AVN and impairment in osteogenic cell compartment following autologous (auto) and allogeneic (allo) HSCT, through the retrospective bone damage revaluation of 100 (50 auto- and 50 allo-HSCT) long-term survivors up to 15 years after transplant. Current treatment options for the management of these complications are also outlined. We found that auto- and allo-HSCT recipients show accelerated bone mineral loss and micro-architectural deterioration during the first years after transplant. Bone mass density (BMD) at the lumbar spine, but not at the femur neck, may improve in some patients after HSCT, suggesting more prolonged bone damage in cortical bone. Phalangeal BMD values remained low for even more years, suggesting persistent bone micro-architectural alterations after transplant. The incidence of AVN was higher in allo-HSCT recipients compared to auto-HSCT recipients. Steroid treatment length, but not its cumulative dose was associated with a higher incidence of bone loss. Allo-HSCT recipients affected by chronic graft versus host disease seem to be at greater risk of continuous bone loss and AVN development. Reduced BMD and higher incidence of AVN was partly related to a reduced regenerating capacity of the normal marrow osteogenic cell compartment. Our results suggest that all patients after auto-HSCT and allo-HSCT should be evaluated for their bone status and treated with anti-resorptive therapy as soon as abnormalities are detected. PMID:23905076

Optimality theories of ageing predict that the balance between reproductive effort and somatic maintenance determines the rate of ageing. Laboratory studies find that increased reproductive effort shortens lifespan, but through increased short-term mortality rather than ageing. In contrast, high fecundity in early life is associated with acceleratedsenescence in free-living vertebrates, but these studies are non-experimental. We performed lifelong brood size manipulation in free-living jackdaws. Actuarial senescence--the increase in mortality rate with age--was threefold higher in birds rearing enlarged- compared to reduced broods, confirming a key prediction of the optimality theory of ageing. Our findings contrast with the results of single-year brood size manipulation studies carried out in many species, in which there was no overall discernible manipulation effect on mortality. We suggest that our and previous findings are in agreement with predictions based on the reliability theory of ageing and propose further tests of this proposition.

Cellular and organismal aging are driven in part by the MTOR (mechanistic target of rapamycin) pathway and rapamycin extends life span inC elegans, Drosophila and mice. Herein, we investigated effects of rapamycin on brain aging in OXYS rats. Previously we found, in OXYS rats, an early development of age-associated pathological phenotypes similar to several geriatric disorders in humans, including cerebral dysfunctions. Behavioral alterations as well as learning and memory deficits develop by 3 months. Here we show that rapamycin treatment (0.1 or 0.5 mg/kg as a food mixture daily from the age of 1.5 to 3.5 months) decreased anxiety and improved locomotor and exploratory behavior in OXYS rats. In untreated OXYS rats, MRI revealed an increase of the area of hippocampus, substantial hydrocephalus and 2-fold increased area of the lateral ventricles. Rapamycin treatment prevented these abnormalities, erasing the difference between OXYS and Wister rats (used as control). All untreated OXYS rats showed signs of neurodegeneration, manifested by loci of demyelination. Rapamycin decreased the percentage of animals with demyelination and the number of loci. Levels of Tau and phospho-Tau (T181) were increased in OXYS rats (compared with Wistar). Rapamycin significantly decreased Tau and inhibited its phosphorylation in the hippocampus of OXYS and Wistar rats. Importantly, rapamycin treatment caused a compensatory increase in levels of S6 and correspondingly levels of phospo-S6 in the frontal cortex, indicating that some downstream events were compensatory preserved, explaining the lack of toxicity. We conclude that rapamycin in low chronic doses can suppress brain aging.

Aging is associated with an increase in oxidative stress and inflammation. The aim of this study was to investigate the effect of aging on various physiological parameters related to inflammation in livers obtained from two types of male mice models: Senescence-accelerated prone (SAMP8) and senescence-accelerated-resistant (SAMR1) mice, and to study the influence of the administration of melatonin (1mg/kg/day) for one month on old SAMP8 mice on these parameters. The parameters studied have been the mRNA expression of TNF-α, iNOS, IL-1β, HO-1, HO-2, MCP1, NFkB1, NFkB2, NFkB protein or NKAP and IL-10. All have been measured by real-time reverse transcription polymerase chain reaction RT-PCR. Furthermore we analyzed the protein expression of TNF-α, iNOS, IL-1β, HO-1, HO-2, and IL-10 by Western-blot. Aging increased oxidative stress and inflammation especially in the liver of SAMP8 mice. Treatment with melatonin decreased the mRNA expression of TNF-α, IL-1β, HO (HO-1 and HO-2), iNOS, MCP1, NFκB1, NFκB2 and NKAP in old male mice. The protein expression of TNF-α, IL-1β was also decreased and IL-10 increased with melatonin treatment and no significant differences were observed in the rest of parameters analyzed. The present study showed that aging was related to inflammation in livers obtained from old male senescence prone mice (SAMP8) and old male senescence resistant mice (SAMR1) being the alterations more evident in the former. Exogenous administration of melatonin was able to reduce inflammation.

Chronic stress can induce cognitive impairment. It is unclear whether a higher susceptibility to chronic stress is associated with the progression of pathological brain aging. Senescence-accelerated prone mouse 8 (SAMP8) is a naturally occurring animal model of accelerated brain aging. Senescence-accelerated resistant mouse 1 (SAMR1) is usually used as the normal control. In this study, we examined the effects of chronic restraint stress (CRS) on learning in the Y-maze, hippocampal cell proliferation, and the expression of brain-derived neurotrophic factor (BDNF) in the hippocampus of 4-month-old SAMP8 and SAMR1. The results showed that exposure to CRS impaired learning and hippocampal cell proliferation in SAMP8 and SAMR1 but to a much greater extent in SAMP8. Furthermore, CRS significantly decreased the expression of BDNF protein and mRNA in the hippocampus of SAMP8 and SAMR1. These data indicated that SAMP8 is more sensitive to the deleterious effects of CRS on learning than SAMR1. A greater decrease in hippocampal cell proliferation caused by chronic stress may be part of the underlying mechanism for the more severe learning deficit observed in SAMP8. In addition, our findings suggested a role of BDNF in the stress-induced impairment of learning and hippocampal cell proliferation in both strains.

Aging is a result of damage accumulation, and understanding of the mechanisms of aging requires exploration of the cellular and molecular systems functioning to control damage. Senescence-accelerated mouse prone 10 (SAMP10) has been established as an inbred strain exhibiting accelerated aging with an earlier onset of cognitive impairment due to neurodegeneration than the senescence-resistant control (SAMR1) strain. We hypothesized that tissue-protective responses of glial cells are impaired in SAMP10 mice. We injected kainic acid (KA) to induce hippocampal injury and studied how cytokines were upregulated on Day 3 using 3-month-old SAMP10 and SAMR1 mice. Following microarray-based screening for upregulated genes, we performed real-time RT-PCR and immunohistochemistry. Results indicated well-orchestrated cytokine-mediated glial interactions in the injured hippocampus of SAMR1 mice, in which microglia-derived interferon (IFN)-γ stimulated astrocytes via IFN-γ receptor and thereby induced expression of CXCL10 and macrophage inflammatory protein (MIP)-1α, and activated microglia produced granulocyte-macrophage colony-stimulating factor (GM-CSF) and osteopontin (OPN). OPN was the most strongly upregulated cytokine. CD44, an OPN receptor, was also strongly upregulated in the neuropil, especially on neurons and astrocytes. KA-induced hippocampal upregulation of these cytokines was strikingly reduced in SAMP10 mice compared to SAMR1 mice. On Day 30 after KA injection, SAMP10 but not SAMR1 mice exhibited hippocampal layer atrophy. Since the OPN-CD44 system is essential for neuroprotection and remodeling, these findings highlight the defects of SAMP10 mice in cytokine-mediated neuroprotective glia-neuron interactions, which may be associated with the mechanism underlying the vulnerability of SAMP10 mice to age-related neurodegeneration.

There is a clear link between overweight, gain of white adipose tissue, and diabetes type 2 (T2D). The molecular mechanism of the gain of adipose tissue is linked with the expression of high mobility group protein AT-hook 2 (HMGA2), and recent studies revealed an association with a SNP near HMGA2. In this study, we investigated the gene expression of HMGA2, p14 (Arf) , CDKN1A, and BAX in human abdominal subcutaneous white adipose tissue from 157 patients. We found a significant higher HMGA2 expression in obese individuals than in non-obese patients. Furthermore, the HMGA2 expression in white adipose tissue in patient with type 2 diabetes was significantly higher than in nondiabetic patients. There is an association between the DNA-binding nonhistone protein HMGA2 and the risk of developing T2D that remains mechanistically unexplained so far. Likewise, p14(Arf), an inducer of cellularsenescence, has been associated with the occurrence of T2D. The data of the present study provide evidence that both proteins act within the same network to drive proliferation of adipose tissue stem and precursor cells, senescence, and increased risk of T2D, respectively.

Some morphogenetic and metabolic processes were sensitive to a high atmospheric CO(2) concentration during sunflower primary leaf ontogeny. Young leaves of sunflower plants growing under elevated CO(2) concentration exhibited increased growth, as reflected by the high specific leaf mass referred to as dry weight in young leaves (16 days). The content of photosynthetic pigments decreased with leaf development, especially in plants grown under elevated CO(2) concentrations, suggesting that high CO(2) accelerates chlorophyll degradation, and also possibly leaf senescence. Elevated CO(2) concentration increased the oxidative stress in sunflower plants by increasing H(2)O(2) levels and decreasing activity of antioxidant enzymes such as catalase and ascorbate peroxidase. The loss of plant defenses probably increases the concentration of reactive oxygen species in the chloroplast, decreasing the photosynthetic pigment content as a result. Elevated CO(2) concentration was found to boost photosynthetic CO(2) fixation, especially in young leaves. High CO(2) also increased the starch and soluble sugar contents (glucose and fructose) and the C/N ratio during sunflower primary leaf development. At the beginning of senescence, we observed a strong increase in the hexoses to sucrose ratio that was especially marked at high CO(2) concentration. These results indicate that elevated CO(2) concentration could promote leaf senescence in sunflower plants by affecting the soluble sugar levels, the C/N ratio and the oxidative status during leaf ontogeny. It is likely that systemic signals produced in plants grown with elevated CO(2), lead to early senescence and a higher oxidation state of the cells of these plant leaves.

Purpose: We examined biological responses of human glioma cells to irradiation with carbon ion beams (C-ions). Methods and Materials: A human glioma-derived cell line, NP-2, was irradiated with C-ions. Apoptotic cell nuclei were stained with Hoechst 33342. Induction of autophagy was examined either by staining cells with monodansylcadaverine (MDC) or by Western blotting to detect conversion of microtuble-associated protein light chain 3 (MAP-LC3) (LC3-I) to the membrane-bound form (LC3-II). Cellularsenescence markers including induction of senescence-associated beta-galactosidase (SA-beta-gal) were examined. The mean telomere length of irradiated cells was determined by Southern blot hybridization. Expression of tumor suppressor p53 and cyclin/cyclin-dependent kinase inhibitor p21{sup WAF1/CIP1} in the irradiated cells was analyzed by Western blotting. Results: When NP-2 cells were irradiated with C-ions at 6 Gy, the major population of the cells died of apoptosis and autophagy. The residual fraction of attached cells (<1% of initially irradiated cells) could not form a colony: however, they showed a morphological phenotype consistent with cellularsenescence, that is, enlarged and flattened appearance. The senescent nature of these attached cells was further indicated by staining for SA-beta-gal. The mean telomere length was not changed after irradiation with C-ions. Phosphorylation of p53 at serine 15 as well as the expression of p21{sup WAF1/CIP1} was induced in NP-2 cells after irradiation. Furthermore, we found that irradiation with C-ions induced cellularsenescence in a human glioma cell line lacking functional p53. Conclusions: Irradiation with C-ions induced apoptosis, autophagy, and cellularsenescence in human glioma cells.

Premature cellularsenescence refers to the state of irreversible cell cycle arrest due to DNA damage or other stresses. In this study, CD9 monoclonal antibody (CD9mAb) was successfully conjugated to the surface of PEGylated liposomes for targeted delivery of rapamycin (LR-CD9mAb) to overcome senescence of CD9 receptor-overexpressing cells. LR-CD9mAb has a small particle size (143.3 ± 2.4 nm), narrow size distribution (polydispersity index: 0.220 ± 0.036), and negative zeta potential (-14.6 ± 1.2 mV). The uptake of CD9-targeted liposomes by premature senescent human dermal fibroblasts (HDFs) was higher than that by young HDFs, as displayed by confocal microscopic images. The senescence might not be reversed by treatment with rapamycin; however, the drug promoted cell proliferation and reduced the number of cells that expressed the senescence-associated-β-galactosidase (SA-β-gal). These effects were further confirmed by cell viability, cell cycle, and Western blotting analyses. Moreover, CD9-targeted liposomes showed better anti-senescence activity, in comparison with free rapamycin or the conventional liposomal formulation, suggesting the potential application of this system in further in vivo studies.

Premature cellularsenescence refers to the state of irreversible cell cycle arrest due to DNA damage or other stresses. In this study, CD9 monoclonal antibody (CD9mAb) was successfully conjugated to the surface of PEGylated liposomes for targeted delivery of rapamycin (LR-CD9mAb) to overcome senescence of CD9 receptor-overexpressing cells. LR-CD9mAb has a small particle size (143.3 ± 2.4 nm), narrow size distribution (polydispersity index: 0.220 ± 0.036), and negative zeta potential (‑14.6 ± 1.2 mV). The uptake of CD9-targeted liposomes by premature senescent human dermal fibroblasts (HDFs) was higher than that by young HDFs, as displayed by confocal microscopic images. The senescence might not be reversed by treatment with rapamycin; however, the drug promoted cell proliferation and reduced the number of cells that expressed the senescence-associated-β-galactosidase (SA-β-gal). These effects were further confirmed by cell viability, cell cycle, and Western blotting analyses. Moreover, CD9-targeted liposomes showed better anti-senescence activity, in comparison with free rapamycin or the conventional liposomal formulation, suggesting the potential application of this system in further in vivo studies.

Ionizing radiation-induced cellularsenescence is thought to be caused by nuclear DNA damage that cannot be repaired. However, here we found that radiation induces delayed increase of intracellular oxidative stress after irradiation. We investigated whether the relief of delayed oxidative stress by ascorbic acid would suppress the radiation-induced cellularsenescence in Syrian golden hamster embryo (SHE) cells. We observed that the level of oxidative stress was drastically increased soon after irradiation, then declined to the level in non-irradiated cells, and increased again with a peak on day 3 after irradiation. We found that the inductions of cellularsenescence after X-irradiation were reduced along with suppression of the delayed induction of oxidative stress by treatment with ascorbic acid, but not when oxidative stress occurred immediately after irradiation. Moreover, treatment of ascorbic acid inhibited p53 accumulation at 3 days after irradiation. Our data suggested a delayed increase of intracellular oxidative stress levels plays an important role in the process of radiation-induced cellularsenescence by p53 accumulation.

Aging is a complex phenomenon and its impact is becoming more relevant due to the rising life expectancy and because aging itself is the basis for the development of age-related diseases such as cancer, neurodegenerative diseases and type 2 diabetes. Recent years of scientific research have brought up different theories that attempt to explain the aging process. So far, there is no single theory that fully explains all facets of aging. The damage accumulation theory is one of the most accepted theories due to the large body of evidence found over the years. Damage accumulation is thought to be driven, among others, by oxidative stress. This condition results in an excess attack of oxidants on biomolecules, which lead to damage accumulation over time and contribute to the functional involution of cells, tissues and organisms. If oxidative stress persists, cellularsenescence is a likely outcome and an important hallmark of aging. Therefore, it becomes crucial to understand how senescent cells function and how they contribute to the aging process. This review will cover cellularsenescence features related to the protein pool such as morphological and molecular hallmarks, how oxidative stress promotes protein modifications, how senescent cells cope with them by proteostasis mechanisms, including antioxidant enzymes and proteolytic systems. We will also highlight the nutritional status of senescent cells and aged organisms (including human clinical studies) by exploring trace elements and micronutrients and on their importance to develop strategies that might increase both, life and health span and postpone aging onset.

Lidamycin (LDM) is a novel member of the enediyne antibiotics identified in China with potent antitumor activity. However, it remains unclear whether LDM has potential molecular targets that may affect its antitumor activity. Enhancer of zeste homolog 2 (EZH2) functions as a histone lysine methyltransferase and mediates trimethylation on histone 3 lysine 27 (H3K27me3). High EZH2 level is found to be positively correlated with the aggressiveness, metastasis and poor prognosis of cancer. Here, we aim to study the role of EZH2 in LDM-induced senescence, as well as in the cytotoxicity of LDM in human colon cancer cells. LDM is found to be relatively more potent in inhibiting the colon cancer cells harboring high EZH2 level and induces irreversible cellularsenescence at IC50 dose range, as evidenced by senescence-associated β-galactosidase staining, cell cycle arrest and molecular changes of senescence regulators including p21 in HCT116 and SW620 cells. More importantly, LDM is found to markedly inhibit EZH2 expression at both protein and mRNA levels upon the induction of p21 and cellularsenescence. LDM also selectively inhibits EZH2 expression as compared with other histone lysine methyltransferases. Knockdown of p21 with siRNAs abolishes LDM-induced senescence, whereas EZH2 knockdown markedly increases p21 expression and causes senescent phenotype. Enrichment of both EZH2 and H3K27me3 levels in the p21 promoter region is reduced by LDM. Moreover, EZH2 overexpression reduces cellularsenescence, p21 expression and DNA damage response upon LDM exposure. LDM also demonstrates potent antitumor efficacy in xenografted animal models. Collectively, our work provides first demonstration that EZH2 may mediate, at least partially, the senescence-inducing effects of LDM by regulating p21 expression and DNA damage effect. Thus, EZH2 may serve as a potential target and biomarker to indicate the clinical efficacy of the potent enediyne antitumor drug. PMID:27882937

Intrahepatic cholestasis of pregnancy (ICP) is a pregnancy-specific disorder characterised by raised bile acids in foetal-maternal circulation, which threatens perinatal health. During the progression of ICP, the effect of oxidative stress is underscored. Peroxiredoxin-3 (PRDX3) is a mitochondrial antioxidant enzyme that is crucial to balance intracellular oxidative stress. However, the role of PRDX3 in placental trophoblast cells under ICP is not fully understood. We demonstrated that the level of PRDX3 was downregulated in ICP placentas as well as bile acids–treated trophoblast cells and villous explant in vitro. Toxic levels of bile acids and PRDX3 knockdown induced oxidative stress and mitochondrial dysfunction in trophoblast cells. Moreover, silencing of PRDX3 in trophoblast cell line HTR8/SVneo induced growth arrest and cellularsenescence via activation of p38-mitogen-activated protein kinase (MAPK) and induction of p21WAF1/CIP and p16INK4A. Additionally, enhanced cellularsenescence, determined by senescence-associated beta-galactosidase staining, was obviously attenuated by p38-MAPK inhibitor SB203580. Our data determined that exposure to bile acid decreased PRDX3 level in human trophoblasts. PRDX3 protected trophoblast cells against mitochondrial dysfunction and cellularsenescence induced by oxidative stress. Our results suggest that decreased PRDX3 by excessive bile acids in trophoblasts plays a critical role in the pathogenesis and progression of ICP. PMID:27958341

Background MER receptor tyrosine kinase (MERTK) is expressed in a variety of malignancies, including glioblastoma multiforme (GBM). Our previous work demonstrated that inhibition of MERTK using RNA interference induced cell death and chemosensitivity in GBM cells, implicating MERTK as a potential therapeutic target. Here we investigate whether a novel MERTK-selective small molecule tyrosine kinase inhibitor, UNC2025, has similar anti-tumor effects in GBM cell lines. Methods Correlations between expression of GAS6, a MERTK ligand, and prognosis were determined using data from the TCGA database. GBM cell lines (A172, SF188, U251) were treated in vitro with increasing doses of UNC2025 (50-400nM). Cell count and viability were determined by trypan blue exclusion. Cell cycle profiles and induction of apoptosis were assessed by flow cytometric analysis after BrdU or Po-Pro-1/propidium iodide staining, respectively. Polyploidy was detected by propidium iodide staining and metaphase spread. Cellularsenescence was determined by β-galactosidase staining and senescence-associated secretory cytokine analysis. Results Decreased overall survival significantly correlated with high levels of GAS6 expression in GBM, highlighting the importance of TAM kinase signaling in GBM tumorigenesis and/or therapy resistance and providing strong rationale for targeting these pathways in the clinic. All three GBM cell lines exhibited dose dependent reductions in cell number and colony formation (>90% at 200nM) after treatment with UNC2025. Cell cycle analysis demonstrated accumulation of cells in the G2/M phase and development of polyploidy. After extended exposure, 60–80% of cells underwent apoptosis. The majority of surviving cells (65–95%) were senescent and did not recover after drug removal. Thus, UNC2025 mediates anti-tumor activity in GBM by multiple mechanisms. Conclusions The findings described here provide further evidence of oncogenic roles for MERTK in GBM, demonstrate the

Several researchers have recently used C6 cells to evaluate functional properties of high-affinity glutamate transporters. However, it has been demonstrated that this lineage suffers several morphological and biochemical alterations according to the number of passages in culture. Currently, there are no reports showing whether functional properties of high-affinity glutamate transporters comply with these sub culturing-dependent modifications. The present study aimed to compare the functional properties of high-affinity glutamate transporters expressed in early (EPC6) and late (LPC6) passage C6 cells through a detailed pharmacological and biochemical characterization. Between 60-180 min of L-[(3)H]glu incubation, LPC6 presented an intracellular [(3)H] 55% lower than EPC6. Both cultures showed a time-dependent increase of intracellular [(3)H] reaching maximal levels at 120 min. Cultures incubated with D-[(3)H]asp showed a time-dependent increase of [(3)H] until 180 min. Moreover, LPC6 have a D-[(3)H]asp-derived intracellular [(3)H] 30-45% lower than EPC6 until 120 min. Only EAAT3 was immunodetected in cultures and its total content was equal between them. PMA-stimulated EAAT3 trafficking to membrane increased 50% of L-[(3)H]glu-derived intracellular [(3)H] in EPC6 and had no effect in LPC6. LPC6 displayed characteristics that resemble senescence, such as high β-Gal staining, cell enlargement and increase of large and regular nuclei. Our results demonstrated that LPC6 exhibited glutamate uptake impairment, which may have occurred due to its inability to mobilize EAAT3 to cell membrane. This profile might be related to senescent process observed in this culture. Our results suggest that LPC6 cells are an inappropriate glial cellular model to investigate the functional properties of high-affinity glutamate transporters.

The mechanisms underlying the development of aging-induced muscle atrophy are unclear. By microRNA array and individual qPCR analyses, we found significant up-regulation of miR-29 in muscles of aged rodents vs. results in young. With aging, p85α, IGF-1 and B-myb muscle levels were lower while the expression of certain cell arrest proteins (p53, p16 and pRB) increased. When miR-29 was expressed in muscle progenitor cells (MPC), their proliferation was impaired while SA-βgal expression increased signifying the development of senescence. Impaired MPC proliferation resulted from interactions between miR-29 and the 3'-UTR of p85a, IGF-1 and B-myb, suppressing the translation of these mediators of myoblast proliferation. In vivo, electroporation of miR-29 into muscles of young mice suppressed the proliferation and increased levels of cellular arrest proteins, recapitulating aging-induced responses in muscle. A potential stimulus of miR-29 expression is Wnt-3a since we found that exogenous Wnt-3a stimulated miR-29 expression 2.7-fold in primary cultures of MPCs. Thus, aging-induced muscle senescence results from activation of miR-29 by Wnt-3a leading to suppressed expression of several signaling proteins (p85α, IGF-1 and B-myb) that act coordinately to impair the proliferation of MPCs contributing to muscle atrophy. The increase in miR-29 provides a potential mechanism for aging-induced sarcopenia.

The mechanisms underlying the development of aging-induced muscle atrophy are unclear. By microRNA array and individual qPCR analyses, we found significant up-regulation of miR-29 in muscles of aged rodents vs. results in young. With aging, p85α, IGF-1 and B-myb muscle levels were lower while the expression of certain cell arrest proteins (p53, p16 and pRB) increased. When miR-29 was expressed in muscle progenitor cells (MPC), their proliferation was impaired while SA-βgal expression increased signifying the development of senescence. Impaired MPC proliferation resulted from interactions between miR-29 and the 3'-UTR of p85a, IGF-1 and B-myb, suppressing the translation of these mediators of myoblast proliferation. In vivo, electroporation of miR-29 into muscles of young mice suppressed the proliferation and increased levels of cellular arrest proteins, recapitulating aging-induced responses in muscle. A potential stimulus of miR-29 expression is Wnt-3a since we found that exogenous Wnt-3a stimulated miR-29 expression 2.7-fold in primary cultures of MPCs. Thus, aging-induced muscle senescence results from activation of miR-29 by Wnt-3a leading to suppressed expression of several signaling proteins (p85α, IGF-1 and B-myb) that act coordinately to impair the proliferation of MPCs contributing to muscle atrophy. The increase in miR-29 provides a potential mechanism for aging-induced sarcopenia. PMID:24659628

Ultraviolet (UV) irradiation generates reactive oxygen species (ROS) in the cells, which induces the cellularsenescence and photoaging. The present study investigated the protective effects of garlic on photo-damage and cellularsenescence in UVB-exposed human keratinocytes, HaCaT cells. An in vitro cell free system was used to examine the scavenging activity of 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals and nitric oxide (NO). The effect of garlic extract on ROS formation, MMP-1 protein and mRNA expressions, cytokines such as interleukin (IL)-1β and IL-6, senescence associated-β-galactosidase (SA-β-gal) activity, and silent information regulator T1 (SIRT1) activity were determined in UVB-irradiated HaCaT cells. Garlic exhibited strong DPPH radical and NO scavenging activity in cell free system exhibiting IC50 values of 2.50 mg/mL and 4.38 mg/mL, respectively. Garlic pretreatment attenuated the production of UVB-induced intracellular ROS. MMP-1 level, which has been known to be induced by ROS, was dramatically elevated by UVB irradiation, and UVB-induced MMP-1 mRNA and protein expressions were significantly reduced by garlic treatment (50 µg/mL) comparable to those of UV-unexposed control cells. UV-induced pro-inflammatory cytokine productions (IL-6 and IL-1β) were significantly inhibited by pretreatment with garlic in a dose-dependent manner. SA-β-gal activity, a classical biomarker of cellularsenescence, and SIRT1 activity, which has attracted attention as an anti-aging factor in recent years, were ameliorated by garlic treatment in UV-irradiated HaCaT cells. The present study provides the first evidence of garlic inhibiting UVB-induced photoaging as a result of augmentation of cellularsenescence in HaCaT human keratinocytes.

Ultraviolet (UV) irradiation generates reactive oxygen species (ROS) in the cells, which induces the cellularsenescence and photoaging. The present study investigated the protective effects of garlic on photo-damage and cellularsenescence in UVB-exposed human keratinocytes, HaCaT cells. An in vitro cell free system was used to examine the scavenging activity of 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals and nitric oxide (NO). The effect of garlic extract on ROS formation, MMP-1 protein and mRNA expressions, cytokines such as interleukin (IL)-1β and IL-6, senescence associated-β-galactosidase (SA-β-gal) activity, and silent information regulator T1 (SIRT1) activity were determined in UVB-irradiated HaCaT cells. Garlic exhibited strong DPPH radical and NO scavenging activity in cell free system exhibiting IC50 values of 2.50 mg/mL and 4.38 mg/mL, respectively. Garlic pretreatment attenuated the production of UVB-induced intracellular ROS. MMP-1 level, which has been known to be induced by ROS, was dramatically elevated by UVB irradiation, and UVB-induced MMP-1 mRNA and protein expressions were significantly reduced by garlic treatment (50 µg/mL) comparable to those of UV-unexposed control cells. UV-induced pro-inflammatory cytokine productions (IL-6 and IL-1β) were significantly inhibited by pretreatment with garlic in a dose-dependent manner. SA-β-gal activity, a classical biomarker of cellularsenescence, and SIRT1 activity, which has attracted attention as an anti-aging factor in recent years, were ameliorated by garlic treatment in UV-irradiated HaCaT cells. The present study provides the first evidence of garlic inhibiting UVB-induced photoaging as a result of augmentation of cellularsenescence in HaCaT human keratinocytes. PMID:27483310

Carbamylated low-density lipoprotein (cLDL) plays a role in atherosclerosis. In this study we evaluate the effect of uremia on LDL carbamylation and the effect of cLDL and oxidized LDL (oxLDL; 200 μg/ml) on number, function, and genomic stability of endothelial progenitor cells (EPCs) obtained from healthy volunteers. cLDL was generated after incubation of native LDL (nLDL) with uremic serum from patients with chronic kidney disease (CKD) stages 2-4. Oxidative stress was measured by flow cytometry and fluorescent microscopy, mitochondrial depolarization by flow cytometry, senescence by β-galactosidase activity and telomere length, and DNA damage by phosphorylated histone H2AX (γH2AX). The percentage of cLDL by uremic serum was related to the severity of CKD. Compared with nLDL, cLDL induced an increase in oxidative stress (62±5 vs. 8±3%, P<0.001) and cells with mitochondrial depolarization (73±7 vs. 9±5%, P<0.001), and a decrease in EPC proliferation and angiogenesis. cLDL also induced acceleratedsenescence (73±16 vs. 12±9%, P<0.001), which was associated with a decrease in the expression of γH2AX (62±9 vs. 5±3%, P<0.001). The degree of injury induced by cLDL was comparable to that observed with oxLDL. This study supports the hypothesis that cLDL triggers genomic damage in EPCs, resulting in premature senescence. We can, therefore, hypothesize that EPCs injury by cLDL contributes to an increase in atherosclerotic disease in CKD.

TP53 is a critical tumor suppressor gene that regulates cell cycle progression, apoptosis, cellularsenescence and many other properties critical for control of normal cellular growth and death. Due to the pleiotropic effects that TP53 has on gene expression and cellular physiology, mutations at this tumor suppressor gene result in diverse physiological effects. T53 mutations are frequently detected in numerous cancers. The expression of TP53 can be induced by various agents used to treat cancer patients such as chemotherapeutic drugs and ionizing radiation. Radiation will induce Ataxia telangiectasia mutated (ATM) and other kinases that results in the phosphorylation and activation of TP53. TP53 is also negatively regulated by other mechanisms, such as ubiquitination by ligases such as MDM2. While TP53 has been documented to control the expression of many "classical" genes (e.g., p21(Cip-1), PUMA, Bax) by transcriptional mechanisms for quite some time, more recently TP53 has been shown to regulate microRNA (miR) gene expression. Different miRs can promote oncogenesis (oncomiR) whereas others act to inhibit tumor progression (tumor suppressor miRs). Targeted therapies to stabilize TP53 have been developed by various approaches, MDM2/MDM4 inhibitors have been developed to stabilize TP53 in TP53-wild type (WT) tumors. In addition, small molecules have been isolated that will reactivate certain mutant TP53s. Both of these types of inhibitors are in clinical trials. Understanding the actions of TP53 may yield novel approaches to suppress cancer, aging and other health problems.

Daphnia (water fleas) are small crustaceans that undergo an unusual switch from asexual to sexual reproduction that is dependent on environmental conditions. In this study, a senescence-associated protein (SAP) from the common freshwater species Daphnia pulex was cloned using primers based on homologous sequences and rapid amplification of cDNA ends (RACE). Real-time PCR was employed to quantify the expression of D. pulex SAP (DpSAP) in individual organisms. The role of DpSAP in the reproductive transformation was further investigated in both parthenogenetic and sexual females by using digoxin-labeled SAP RNA probes and RNA whole-mount in situ hybridization. DpSAP was more highly expressed in sexual females, indicating a role in growth and reproduction. Cellular localization studies using RNA whole-mount in situ hybridization showed specific expression in the second tentacle joints. These expression patterns suggest an important role for DpSAP in the reproductive transformation of D. pulex.

Human mesenchymal stem cells (hMSCs) have great therapeutic potential due to their high plasticity, immune privileged status and ease of preparation, as well as a lack of ethical barriers to their use. However, their ultimate usefulness is limited by cellularsenescence occurring secondary to increased cellular levels of reactive oxygen species (ROS) during their propagation in culture. The underlying molecular mechanisms responsible for this process in hMSCs remain unclear. An antioxidant polyphenol epigallocatechin-3-gallate (EGCG) found in green tea, is known to activate nuclear factor-erythroid 2-related factor 2 (Nrf2), a master transcriptional regulator of antioxidant genes. Herein, we examined the EGCG-mediated antioxidant mechanism in hMSCs exposed to ROS which involves Nrf2 activation. The H2O2-exposed hMSCs showed cellularsenescence with significantly increased protein levels of acetyl-p53 and p21 in comparison with the untreated hMSCs, and these effects were prevented by pre-treatment with EGCG. By contrast, in Nrf2-knockdown hMSCs, EGCG lost its antioxidant effect, exhibiting high levels of acetyl-p53 and p21 following EGCG pre-treatment and H2O2 exposure. This indicates that Nrf2 and p53/p21 may be involved in the anti‑senescent effect of EGCG in hMSCs. Taken together, these findings indicate the important role of EGCG in preventing oxidative stress-induced cellularsenescence in hMSCs through Nrf2 activation, which has applications for the massive production of more suitable hMSCs for cell-based therapy.

Hyperactivation of the Wingless-type (Wnt)/β-catenin pathway promotes tumor initiation, tumor growth and metastasis in various tissues. Although there is evidence for the involvement of Wnt/β-catenin pathway activation in salivary gland tumors, the precise mechanisms are unknown. Here we report for the first time that downregulation of the Wnt inhibitory factor 1 (WIF1) is a widespread event in salivary gland carcinoma ex-pleomorphic adenoma (CaExPA). We also show that WIF1 downregulation occurs in the CaExPA precursor lesion pleomorphic adenoma (PA) and indicates a higher risk of progression from benign to malignant tumor. Our results demonstrate that diverse mechanisms including WIF1 promoter hypermethylation and loss of heterozygosity contribute to WIF1 downregulation in human salivary gland tumors. In accordance with a crucial role in suppressing salivary gland tumor progression, WIF1 re-expression in salivary gland tumor cells inhibited cell proliferation, induced more differentiated phenotype and promoted cellularsenescence, possibly through upregulation of tumor-suppressor genes, such as p53 and p21. Most importantly, WIF1 significantly diminished the number of salivary gland cancer stem cells and the anchorage-independent cell growth. Consistent with this observation, WIF1 caused a reduction in the expression of pluripotency and stemness markers (OCT4 and c-MYC), as well as adult stem cell self-renewal and multi-lineage differentiation markers, such as WNT3A, TCF4, c-KIT and MYB. Furthermore, WIF1 significantly increased the expression of microRNAs pri-let-7a and pri-miR-200c, negative regulators of stemness and cancer progression. In addition, we show that WIF1 functions as a positive regulator of miR-200c, leading to downregulation of BMI1, ZEB1 and ZEB2, with a consequent increase in downstream targets such as E-cadherin. Our study emphasizes the prognostic and therapeutic potential of WIF1 in human salivary gland CaExPA. Moreover, our findings

DNA-damage induces a DNA-damage response (DDR) in mammalian cells. The response, depending upon the cell-type and the extent of DNA-damage, ultimately results in cell death or cellularsenescence. DDR-induced signaling in cells activates the ATM-p53 and ATM-IKKα/β-interferon (IFN)-β signaling pathways, thus leading to an induction of the p53 and IFN-inducible IFI16 gene. Further, upon DNA-damage, DNA accumulates in the cytoplasm, thereby inducing the IFI16 protein and STING-dependent IFN-β production and activation of the IFI16 inflammasome, resulting in the production of proinflammatory cytokines (e.g., IL-1β and IL-18). Increased expression of IFI16 protein in a variety of cell-types promotes cellularsenescence. However, reduced expression of IFI16 in cells promotes cell proliferation. Because expression of the IFI16 gene is induced by activation of DNA-damage response in cells and increased levels of IFI16 protein in cells potentiate the p53-mediated transcriptional activation of genes and p53 and pRb-mediated cell cycle arrest, we discuss how an improved understanding of the role of IFI16 protein in cellularsenescence and associated inflammatory secretory phenotype is likely to identify the molecular mechanisms that contribute to the development of aging-associated human inflammatory diseases and a failure to cancer therapy.

Senescence is a form of cell cycle arrest induced by stress such as DNA damage and oncogenes. However, while arrested, senescent cells secrete a variety of proteins collectively known as the senescence-associated secretory phenotype (SASP), which can reinforce the arrest and induce senescence in a paracrine manner. However, the SASP has also been shown to favor embryonic development, wound healing, and even tumor growth, suggesting more complex physiological roles than currently understood. Here we uncover timely new functions of the SASP in promoting a proregenerative response through the induction of cell plasticity and stemness. We show that primary mouse keratinocytes transiently exposed to the SASP exhibit increased expression of stem cell markers and regenerative capacity in vivo. However, prolonged exposure to the SASP causes a subsequent cell-intrinsic senescence arrest to counter the continued regenerative stimuli. Finally, by inducing senescence in single cells in vivo in the liver, we demonstrate that this activates tissue-specific expression of stem cell markers. Together, this work uncovers a primary and beneficial role for the SASP in promoting cell plasticity and tissue regeneration and introduces the concept that transient therapeutic delivery of senescent cells could be harnessed to drive tissue regeneration. PMID:28143833

Oncogene-induced senescence (OIS) is a critical tumor-suppressing mechanism that restrains cancer progression at premalignant stages, in part by causing telomere dysfunction. Currently it is unknown whether this proliferative arrest presents a stable and therefore irreversible barrier to cancer progression. Here we demonstrate that cells frequently escape OIS induced by oncogenic H-Ras and B-Raf, after a prolonged period in the senescence arrested state. Cells that had escaped senescence displayed high oncogene expression levels, retained functional DNA damage responses, and acquired chromatin changes that promoted c-Myc–dependent expression of the human telomerase reverse transcriptase gene (hTERT). Telomerase was able to resolve existing telomeric DNA damage response foci and suppressed formation of new ones that were generated as a consequence of DNA replication stress and oncogenic signals. Inhibition of MAP kinase signaling, suppressing c-Myc expression, or inhibiting telomerase activity, caused telomere dysfunction and proliferative defects in cells that had escaped senescence, whereas ectopic expression of hTERT facilitated OIS escape. In human early neoplastic skin and breast tissue, hTERT expression was detected in cells that displayed features of senescence, suggesting that reactivation of telomerase expression in senescent cells is an early event during cancer progression in humans. Together, our data demonstrate that cells arrested in OIS retain the potential to escape senescence by mechanisms that involve derepression of hTERT expression. PMID:27503890

Hodgkin's Lymphoma (HL) is one of the most prevailing malignancies in young adults. Reed–Sternberg (RS) cells in HL have distinctive large cell morphology, are characteristic of the disease and their presence is essential for diagnosis. Enlarged cells are one of the hallmarks of senescence, but whether RS cells are senescent has not been previously investigated. Here we show that RS cells have characteristics of senescent cells; RS cells in HL biopsies specifically express the senescence markers and cell cycle inhibitors p21Cip1 and p16INK4a and are negative for the proliferation marker Ki-67, suggesting that these cells have ceased to proliferate. Moreover, the RS-like cells in HL lines, stained specifically for senescence-associated β-galactosidase (SA-β-gal). Oxidative stress promoted senescence in these cells as demonstrated by their staining for p21Cip1, p16INK4a, p53 and γH2AX. Senescent cells produce copious amounts of inflammatory cytokines termed ‘senescence-associated secretory phenotype' (SASP), primarily regulated by Nuclear Factor κB (NF-κB). Indeed, we show that NF-κB activity and NF-κB-dependent cytokines production (e.g., IL-6, TNF-α, GM-CSF) were elevated in RS-like cells. Furthermore, NF-κB inhibitors, JSH-23 and curcumin reduced IL-6 secretion from RS-like cells. Thus, defining RS cells as senescent offers new insights on the origin of the proinflammatory microenvironment in HL. PMID:27831553

AKTIP is a shelterin-interacting protein required for replication of telomeric DNA. Here, we show that AKTIP biochemically interacts with A- and B-type lamins and affects lamin A, but not lamin C or B, expression. In interphase cells, AKTIP localizes at the nuclear rim and in discrete regions of the nucleoplasm just like lamins. Double immunostaining revealed that AKTIP partially co-localizes with lamin B1 and lamin A/C in interphase cells, and that proper AKTIP localization requires functional lamin A. In mitotic cells, AKTIP is enriched at the spindle poles and at the midbody of late telophase cells similar to lamin B1. AKTIP-depleted cells show senescence-associated markers and recapitulate several aspects of the progeroid phenotype. Collectively, our results indicate that AKTIP is a new player in lamin-related processes, including those that govern nuclear architecture, telomere homeostasis and cellularsenescence. PMID:27512140

AKTIP is a shelterin-interacting protein required for replication of telomeric DNA. Here, we show that AKTIP biochemically interacts with A- and B-type lamins and affects lamin A, but not lamin C or B, expression. In interphase cells, AKTIP localizes at the nuclear rim and in discrete regions of the nucleoplasm just like lamins. Double immunostaining revealed that AKTIP partially co-localizes with lamin B1 and lamin A/C in interphase cells, and that proper AKTIP localization requires functional lamin A. In mitotic cells, AKTIP is enriched at the spindle poles and at the midbody of late telophase cells similar to lamin B1. AKTIP-depleted cells show senescence-associated markers and recapitulate several aspects of the progeroid phenotype. Collectively, our results indicate that AKTIP is a new player in lamin-related processes, including those that govern nuclear architecture, telomere homeostasis and cellularsenescence.

Recent experiments have shown that cellularsenescence, a mechanism employed by cells for thwarting cell proliferation, plays an important role in protecting cells against cancer; therefore, a deeper understanding of cellularsenescence can lead to effective cancer treatment. Inhibition of CDK2 is thought to be the critical trigger for cellularsenescence. In this study, we first implement a mathematical model of G1/S transition involving the DNA-damage pathway and show that cellularsenescence can be achieved by lowering CDK2. The robustness of CDK2 in triggering cellularsenescence is determined from the probability (β) of DNA-damaged cells passing G1/S checkpoint for normal CDK2 and CDK2-deficient situations based on different thresholds of the peak time of two important biomarkers, CycE and E2F. The comparison of the values of β under the normal CDK2 and lower CDK2 levels reveals that reducing CDK2 levels can decrease the percentage of damaged cells passing G1/S checkpoint; more importantly, 50% reduction of CDK2 achieves 65% reduction in the percentage of damaged cells passing the G1/S checkpoint. These results point out that the developed model can highlight the possibility of lowering the bar for cellularsenescence by reducing CDK2 levels. The results of investigation of β for the different thresholds of the peak times of other biomarkers show that β is insensitive to these perturbations of the peak time indicating that CDK2 activity is robust in lowering the senescence bar for low and high levels of DNA-damage. Furthermore, a mathematical formulation of robustness indicates that the robustness of CDK2-triggered senescence increases with decreasing levels of CDK2, and is slightly greater for low-level DNA damage condition.

TopBP1 plays important roles in chromosome replication, DNA damage response, and other cellular regulatory functions in vertebrates. Although the roles of TopBP1 have been studied mostly in cancer cell lines, its physiological function remains unclear in mice and untransformed cells. We generated conditional knock-out mice in which exons 5 and 6 of the TopBP1 gene are flanked by loxP sequences. Although TopBP1-deficient embryos developed to the blastocyst stage, no homozygous mutant embryos were recovered at E8.5 or beyond, and completely resorbed embryos were frequent at E7.5, indicating that mutant embryos tend to die at the peri-implantation stage. This finding indicated that TopBP1 is essential for cell proliferation during early embryogenesis. Ablation of TopBP1 in TopBP1(flox/flox) mouse embryonic fibroblasts and 3T3 cells using Cre recombinase-expressing retrovirus arrests cell cycle progression at the G(1), S, and G(2)/M phases. The TopBP1-ablated mouse cells exhibit phosphorylation of H2AX and Chk2, indicating that the cells contain DNA breaks. The TopBP1-ablated mouse cells enter cellularsenescence. Although RNA interference-mediated knockdown of TopBP1 induced cellularsenescence in human primary cells, it induced apoptosis in cancer cells. Therefore, TopBP1 deficiency in untransformed mouse and human primary cells induces cellularsenescence rather than apoptosis. These results indicate that TopBP1 is essential for cell proliferation and maintenance of chromosomal integrity.

Dietary interventions have been proposed as a way to increase lifespan and improve health. The senescence-accelerated prone 8 (SAMP8) mice have a shorter lifespan and show alterations in the central nervous system. Moreover, this mouse strain shows decreased sirtuin 1 protein expression and elevated expression of the acetylated targets NFkappaB and FoxO1, which are implicated in transcriptional control of key genes in cell proliferation and cell survival, in reference to control strain, SAMR1. After eight weeks of intermittent fasting, sirtuin 1 protein expression was recovered in SAMP8. This recovery was accompanied by a reduction in the two acetylated targets. Furthermore, SAMP8 showed a lower protein expression of BDNF and HSP70 while intermittent fasting re-established normal values. The activation of JNK and FoxO1 was also reduced in SAMP8 mice subjected to an IF regimen, compared with control SAMP8. Our findings provide new insights into the participation of sirtuin 1 in ageing and point to a potential novel application of this enzyme to prevent frailty due to ageing processes in the brain.

The histidine-containing dipeptides (HCD) carnosine and anserine are found in high concentrations in mammalian skeletal muscle. Given its versatile biologic properties, such as antioxidative, antiglycation, and pH buffering capacity, carnosine has been implicated as a protective factor in the aging process. The present study aimed to systematically explore age-related changes in skeletal muscles HCD content in a murine model of accelerated aging. Additionally, we investigated the effect of lifelong creatine supplementation on muscle HCD content and contractile fatiguability. Male senescence-accelerated mice (SAMP8) were fed control or creatine-supplemented (2% of food intake) diet from the age of 10 to 60 weeks. At week 10, 25, and 60, tibialis anterior muscles were dissected and analysed for HCD and taurine content by HPLC. Soleus and EDL muscles were tested for in vitro contractile fatigue and recovery. From 10 to 60 weeks of age, muscular carnosine (-45%), taurine (-24%), and total creatine (-42%) concentrations gradually and significantly decreased. At 25 but not at 60 weeks, oral creatine supplementation significantly increased carnosine (+88%) and anserine (+40%) content compared to age-matched control-fed animals. Taurine and total creatine content were not affected by creatine supplementation at any age. Creatine-treated mice showed attenuated muscle fatigue (soleus) and enhanced force recovery (m. extensor digitorum longus [EDL]) compared to controls at 25 weeks, but not at 60 weeks. From the present study, we can conclude that skeletal muscle tissue exhibits a significant decline in HCD content at old age. Oral creatine supplementation is able to transiently but potently increase muscle carnosine and anserine content, which coincides with improved resistance to contractile fatigue.

Summary The RB protein family (RB, p107, p130) has overlapping and compensatory functions in cell cycle control. However, cancer-associated mutations are almost exclusively found in RB, implying that RB has a non-redundant role in tumor suppression. We demonstrate that RB preferentially associates with E2F target genes involved in DNA replication and is uniquely required to repress these genes during senescence but not other growth states. Consequently, RB loss leads to inappropriate DNA synthesis following a senescence trigger and, together with disruption of a p21-mediated cell cycle checkpoint, enables extensive proliferation and rampant genomic instability. Our results identify a non-redundant RB effector function that may contribute to tumor suppression and reveal how loss of RB and p53 cooperate to bypass senescence. PMID:20385362

The RB protein family (RB, p107, and p130) has overlapping and compensatory functions in cell-cycle control. However, cancer-associated mutations are almost exclusively found in RB, implying that RB has a nonredundant role in tumor suppression. We demonstrate that RB preferentially associates with E2F target genes involved in DNA replication and is uniquely required to repress these genes during senescence but not other growth states. Consequently, RB loss leads to inappropriate DNA synthesis following a senescence trigger and, together with disruption of a p21-mediated cell-cycle checkpoint, enables extensive proliferation and rampant genomic instability. Our results identify a nonredundant RB effector function that may contribute to tumor suppression and reveal how loss of RB and p53 cooperate to bypass senescence.

Petal development and senescence entails a normally irreversible process. It starts with petal expansion and pigment production, and ends with nutrient remobilization and ultimately cell death. In many species this is accompanied by petal abscission. Post-harvest stress is an important factor in limiting petal longevity in cut flowers and accelerates some of the processes of senescence such as petal wilting and abscission. However, some of the effects of moderate stress in young flowers are reversible with appropriate treatments. Transcriptomic studies have shown that distinct gene sets are expressed during petal development and senescence. Despite this, the overlap in gene expression between developmental and stress-induced senescence in petals has not been fully investigated in any species. Here a custom-made cDNA microarray from Alstroemeria petals was used to investigate the overlap in gene expression between developmental changes (bud to first sign of senescence) and typical post-harvest stress treatments. Young flowers were stressed by cold or ambient temperatures without water followed by a recovery and rehydration period. Stressed flowers were still at the bud stage after stress treatments. Microarray analysis showed that ambient dehydration stress accelerates many of the changes in gene expression patterns that would normally occur during developmental senescence. However, a higher proportion of gene expression changes in response to cold stress were specific to this stimulus and not senescence related. The expression of 21 transcription factors was characterized, showing that overlapping sets of regulatory genes are activated during developmental senescence and by different stresses.

Pneumonia generates considerable negative impacts on the elderly. Despite the widespread uses of vaccines and appropriate antibiotics, the morbidity and mortality of elderly pneumonia are significantly higher compared to the counterparts of young populations. The definitive mechanisms of high vulnerability in the elderly against pathogen threats are unclear. Age-associated, chronic low-grade inflammation augments the susceptibility and severity of pneumonia in the elderly. Cellularsenescence, one of the hallmarks of aging, has its own characteristics, cell growth arrest and senescence-associated secretory phenotype (SASP). These properties are beneficial if the sequence of senescence-clearance-regeneration is transient in manner. However, persisting senescent cell accumulation and excessive SASP might induce sustained low-grade inflammation and disruption of normal tissue microenvironments in aged tissue. Emerging evidence indicates that cellularsenescence is a key component in the pathogenesis of chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF), which are known to be age-related and increase the risk of pneumonia. In addition to their structural collapses, COPD and IPF might increase the vulnerability to pathogen insults through SASP. Here, we discuss the current advances in understanding of the impacts of cellularsenescence in elderly pneumonia and in these chronic lung disorders that heighten the risk of respiratory infections.

When cells are grown to confluence, cell-cell contact inhibition occurs and drives the cells to enter reversible quiescence rather than senescence. Confluent retinal pigment epithelial (RPE) cells exhibiting contact inhibition was used as a model in this study to examine the role of overexpression of transcription factor AP4, a highly expressed transcription factor in many types of cancer, in these cells during long-term culture. We generated stable inducible RPE cell clones expressing AP4 or AP4 without the DNA binding domain (DN-AP4) and observed that, when cultured for 24 days, RPE cells with a high level of AP4 exhibit a large, flattened morphology and even cease proliferating; these changes were not observed in DN-AP4-expressing cells or non-induced cells. In addition, AP4-expressing cells exhibited senescence-associated β-galactosidase activity and the senescence-associated secretory phenotype. We demonstrated that the induced cellularsenescence was mediated by enhanced p53 expression and that AP4 regulates the p53 gene by binding directly to two of the three E-boxes present on the promoter of the p53 gene. Moreover, we showed that serum is essential for AP4 in inducing p53-associated cellularsenescence. Collectively, we showed that overexpression of AP4 mediates cellularsenescence involving in activation of p53 in long-term post-confluent RPE cells.

Although it has been reported that mesenchymal stromal cells are unable to provide sufficient hematopoietic support in myelodysplastic syndrome, the underlying mechanisms remain elusive. In this study, we found that mesenchymal stromal cells from patients with myelodysplastic syndrome displayed a significant increase in senescence, as evidenced by their decreased proliferative capacity, flattened morphology and increased expression of SA-β-gal and p21. Senescent mesenchymal stromal cells from patients had decreased differentiation potential and decreased stem cell support capacity. Gene knockdown of Dicer1, which was down-regulated in mesenchymal stromal cells from patients, induced senescence. The differentiation and stem cell-supporting capacities were significantly inhibited by Dicer1 knockdown. Overexpression of Dicer1 in mesenchymal stromal cells from patients reversed cellularsenescence and enhanced stem cell properties. Furthermore, we identified reduced expression in the microRNA-17 family (miR-17-5p, miR-20a/b, miR-106a/b and miR-93) as a potential factor responsible for increased p21 expression, a key senescence mediator, in Dicer1 knockdown cells. Moreover, we found that miR-93 and miR-20a expression levels were significantly reduced in mesenchymal stromal cells from patients and miR-93/miR-20a gain of function resulted in a decrease of cellularsenescence. Collectively, the results of our study show that mesenchymal stromal cells from patients with myelodysplastic syndrome are prone to senescence and that Dicer1 down-regulation promotes cellularsenescence and decreases the differentiation and stem cell-supporting capacities of mesenchymal stromal cells. Dicer1 down-regulation seems to contribute to the insufficient hematopoietic support capacities of mesenchymal stromal cells from patients with myelodysplastic syndrome.

Cyclin-dependent kinase inhibitor p16INK4a (p16) primarily functions as a negative regulator of the retinoblastoma protein (Rb) -E2F pathway, thus plays critical role in cell cycle progression, cellularsenescence and apoptosis. In this study, we showed that the methylation of Arg 138 and the phosphorylation of Ser 140 on p16 were critical for the control of cell proliferation and apoptosis. Compared to wild type p16, mutant p16R138K possessed improved function in preventing cell proliferation and inducing apoptosis, while the Ser 140 mutation (p16S140A) exhibited the opposite alteration. We also demonstrated that H2O2 was able to induce the phosphorylation of p16, which facilitated the interaction between CDK4 (Cyclin-dependent protein kinase) and p16, in 293T (human emborynic kidney) cells. Furthermore, the elevated arginine methylation in p16S140A mutant and increased serine phosphorylation in p16R138K mutant suggest that a antagonizing mechanism coordinating Arg 138 methylation and Ser 140 phosphorylation to regulates p16 function as well as cellular apoptosis and senescence. These findings will therefore contribute to therapeutic treatment for p16-related gene therapy by providing theoretical and experimental evidence. PMID:28120917

The transcription factor nuclear factor kappa B (NF-κB) and the long non-coding RNA (lncRNA) HOTAIR (HOX transcript antisense RNA) play diverse functional roles in cancer. In this study, we show that upregulation of HOTAIR induced platinum resistance in ovarian cancer, and increased HOTAIR levels were observed in recurrent platinum-resistant ovarian tumors vs. primary ovarian tumors. To investigate the role of HOTAIR during DNA damage induced by platinum, we monitored double-strand breaks and show that HOTAIR expression results in sustained activation of DNA damage response after platinum treatment. We demonstrate that ectopic expression of HOTAIR induces NF-κB activation during DNA damage response and MMP-9 and IL-6 expression, both key NF-κB target genes. We show that HOTAIR regulates activation of NF-κB by decreasing Iκ-Bα (NF-κB inhibitor) and establish that by inducing prolonged NF-κB activation and expression of NF-κB target genes during DNA damage, HOTAIR plays a critical role in cellularsenescence and platinum sensitivity. Our findings suggest that a NF-κB-HOTAIR axis drives a positive-feedback loop cascade during DNA damage response and contributes to cellularsenescence and chemotherapy resistance in ovarian and other cancers. PMID:27041570

Cyclin-dependent kinase inhibitor p16(INK4a) (p16) primarily functions as a negative regulator of the retinoblastoma protein (Rb) -E2F pathway, thus plays critical role in cell cycle progression, cellularsenescence and apoptosis. In this study, we showed that the methylation of Arg 138 and the phosphorylation of Ser 140 on p16 were critical for the control of cell proliferation and apoptosis. Compared to wild type p16, mutant p16R138K possessed improved function in preventing cell proliferation and inducing apoptosis, while the Ser 140 mutation (p16S140A) exhibited the opposite alteration. We also demonstrated that H2O2 was able to induce the phosphorylation of p16, which facilitated the interaction between CDK4 (Cyclin-dependent protein kinase) and p16, in 293T (human emborynic kidney) cells. Furthermore, the elevated arginine methylation in p16S140A mutant and increased serine phosphorylation in p16R138K mutant suggest that a antagonizing mechanism coordinating Arg 138 methylation and Ser 140 phosphorylation to regulates p16 function as well as cellular apoptosis and senescence. These findings will therefore contribute to therapeutic treatment for p16-related gene therapy by providing theoretical and experimental evidence.

Summary We investigated miRNA expression changes associated with stress-induced premature senescence (SIPS) in primary cultures of human diploid fibroblasts (HDF) and human trabecular meshwork (HTM) cells. Twenty-five miRNAs were identified by miRNA microarray analysis and their changes in expression were validated by TaqMan realtime RT-PCR in three independent cell lines of HTM and HDF. SIPS in both HTM and HDF cell types was associated with significant down-regulation of four members of the miR-15 family and five miRNAs of the miR-106b family located in the oncogenic clusters miR-17–92, miR-106a-363, and miR-106b-25. SIPS was also associated with up-regulation of two miRNAs (182 and 183) from the miR-183-96-182 cluster. Transfection with miR-106a agomir inhibited the up-regulation of p21CDKN1A associated with SIPS while transfection with miR-106a antagomir led to increased p21CDKN1A expression in senescent cells. In addition, we identified retinoic acid receptor gamma (RARG) as a target of miR-182 and showed that this protein was down-regulated during SIPS in HDF and HTM cells. These results suggest that changes in miRNA expression might contribute to phenotypic alterations of senescent cells by modulating the expression of key regulatory proteins such as p21CDKN1A as well as by targeting genes that are down-regulated in senescent cells such as RARG. PMID:19782699

Although muscle wasting and/or degeneration are prevalent in patients with chronic kidney disease, it remains unknown whether FGF-23 influences muscle homeostasis and regeneration. Mesenchymal stem cells (MSCs) in skeletal muscle are distinct from satellite cells and have a known association with muscle degeneration. In this study we sought to investigate the effects of FGF-23 on MSCs isolated from human skeletal muscle in vitro. The MSCs expressed FGF receptors (1 through 4) and angiotensin-II type 1 receptor, but no traces of the Klotho gene were detected. MSCs and satellite cells were treated with FGF-23 and angiotensin-II for 48 h. Treatment with FGF-23 significantly decreased the number of MSCs compared to controls, while treatment with angiotensin-II did not. FGF-23 and angiotensin-II both left the cell counts of the satellite cells unchanged. The FGF-23-treated MSCs exhibited the senescent phenotype, as judged by senescence-associated β-galactosidase assay, cell morphology, and increased expression of p53 and p21 in western blot analysis. FGF-23 also significantly altered the gene expression of oxidative stress regulators in the cells. In conclusion, FGF-23 induced premature senescence in MSCs from skeletal muscle via the p53/p21/oxidative-stress pathway. The interaction between the MSCs and FGF-23 may play a key role in the impaired muscle reparative mechanisms of chronic kidney disease.

The human Telomerase Reverse Transcriptase (hTERT) gene encodes a rate-limiting catalytic subunit of telomerase that maintains genomic integrity. Suppression of hTERT expression could induce cellularsenescence and is considered a potent approach for gastric cancer therapy. However, control of hTERT expression and function remains poorly understood in gastric cancer. In this study, we demonstrated that high expression levels of hTERT in malignant tissues are correlated with poor survival probability in gastric cancer patients. Knockdown of hTERT expression retarded cell proliferation and cellularsenescence, which was confirmed by increased protein expression levels of p21cip1 and p27kip1, and decreased phosphorylation of Rb. In contrast, overexpression of hTERT increased cell proliferation and decreased cellularsenescence. Remarkably, the down-regulation of hTERT expression was detected in lgals3−/− mouse embryo fibroblasts (MEFs). Knockdown of galectin-3 decreased the expression of hTERT in gastric cancer cells. Galectin-3 ablation-induced cellularsenescence was rescued by concomitant overexpression of hTERT. hTERT ablation-induced cellularsenescence and p21cip1 and p27kip1 expression was rescued by concomitant overexpression of galectin-3. The size of tumor burdens was increased in hTERT-overexpressed gastric cancer cells xenografted mice, whereas it was repressed by concomitant depletion of galectin-3. Additionally, we determined that the N-terminal domain of galectin-3 directly interacted with hTERT. The telomeric activity of hTERT was also decreased by galectin-3 ablation. Taken together, ablation of hTERT induces cellularsenescence and inhibits the growth of gastric cancer cells, suggesting that it could be a potent target in gastric cancer therapy. We also propose that galectin-3 is an important regulator of hTERT expression and telomeric activity in gastric tumorigenesis. PMID:27494887

Long noncoding RNAs (lncRNAs) may play an important role in Alzheimer's disease (AD) pathogenesis. However, despite considerable research in this area, the comprehensive and systematic understanding of lncRNAs in AD is still limited. The emergence of RNA sequencing provides a predictor and has incomparable advantage compared with other methods, including microarray. In this study, we identified lncRNAs in a 7-month-old mouse brain through deep RNA sequencing using the senescence-accelerated mouse prone 8 (SAMP8) and senescence-accelerated mouse resistant 1 (SAMR1) models. A total of 599,985,802 clean reads and 23,334 lncRNA transcripts were obtained. Then, we identified 97 significantly upregulated and 114 significantly downregulated lncRNA transcripts from all cases in SAMP8 mice relative to SAMR1 mice. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes analyses revealed that these significantly dysregulated lncRNAs were involved in regulating the development of AD from various angles, such as nerve growth factor term (GO: 1990089), mitogen-activated protein kinase signaling pathway, and AD pathway. Furthermore, the most probable AD-associated lncRNAs were predicted and listed in detail. Our study provided the systematic dissection of lncRNA profiling in SAMP8 mouse brain and accelerated the development of lncRNA biomarkers in AD. These attracting biomarkers could provide significant insights into AD therapy in the future. PMID:27483026

In the present investigation we aim to provide experimental information on and thereby understanding of the generation and propagation of spark-ignited, outwardly propagating cellular flames, with three major focuses. The first is to unambiguously demonstrate the influence of the four most important parameters in inducing hydrodynamic and diffusional-thermal cellularities, namely thermal expansion, flame thickness, non-unity Lewis number, and global activation energy. The second is to investigate the critical state for the onset of cellularity for the stretch-affected, expanding flame. The third is to identify and consequently quantify the phenomena of self-acceleration and possibly auto-turbulization of cellular flames. Due to space limitation the effects of activation energy and the critical state for the onset of cellularity will not be discussed herein. Experiments were conducted using C3H8-air and H2-O2-N2 mixtures for their opposite influences of non-equidiffusivity. The additional system parameters varied were the chamber pressure (p) and the mixture composition including the equivalence ratio (phi). From a sequence of the flame images we can assess the propensity of cell formation, and determine the instantaneous flame radius (R), the flame propagation rate, the global stretch rate experienced by the flame, the critical flame radius at which cells start to grow, and the average cell size.

Methylene blue (MB) delays cellularsenescence, induces complex-IV, and activates Keap1/Nrf2; however, the molecular link of these effects to MB is unclear. Since MB is redox-active, we investigated its effect on the NAD/NADH ratio in IMR90 cells. The transient increase in NAD/NADH observed in MB-treated cells triggered an investigation of the energy regulator AMPK. MB induced AMPK phosphorylation in a transient pattern, which was followed by the induction of PGC1α and SURF1: both are inducers of mitochondrial and complex-IV biogenesis. Subsequently MB-treated cells exhibited >100% increase in complex-IV activity and a 28% decline in cellular oxidants. The telomeres erosion rate was also significantly lower in MB-treated cells. A previous research suggested that the pattern of AMPK activation (i.e., chronic or transient) determines the AMPK effect on cell senescence. We identified that the anti-senescence activity of MB (transient activator) was 8-times higher than that of AICAR (chronic activator). Since MB lacked an effect on cell cycle, an MB-dependent change to cell cycle is unlikely to contribute to the anti-senescence activity. The current findings in conjunction with the activation of Keap1/Nrf2 suggest a synchronized activation of the energy and cellular defense pathways as a possible key factor in MB's potent anti-senescence activity.

Pneumonia generates considerable negative impacts on the elderly. Despite the widespread uses of vaccines and appropriate antibiotics, the morbidity and mortality of elderly pneumonia are significantly higher compared to the counterparts of young populations. The definitive mechanisms of high vulnerability in the elderly against pathogen threats are unclear. Age-associated, chronic low-grade inflammation augments the susceptibility and severity of pneumonia in the elderly. Cellularsenescence, one of the hallmarks of aging, has its own characteristics, cell growth arrest and senescence-associated secretory phenotype (SASP). These properties are beneficial if the sequence of senescence–clearance–regeneration is transient in manner. However, persisting senescent cell accumulation and excessive SASP might induce sustained low-grade inflammation and disruption of normal tissue microenvironments in aged tissue. Emerging evidence indicates that cellularsenescence is a key component in the pathogenesis of chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF), which are known to be age-related and increase the risk of pneumonia. In addition to their structural collapses, COPD and IPF might increase the vulnerability to pathogen insults through SASP. Here, we discuss the current advances in understanding of the impacts of cellularsenescence in elderly pneumonia and in these chronic lung disorders that heighten the risk of respiratory infections. PMID:28245616

Telomeres are protective structures at the ends of chromosomes that have important implications for aging. To address the question of whether telomeres contribute to feline chronic kidney disease (CKD), we evaluated kidney, liver, and skin samples from 12 cats with naturally occurring CKD, 12 young normal cats, and 6 old normal cats. Telomere length was assessed using standard telomere fluorescent in situ hybridization (TEL-FISH) combined with immunohistochemistry (TELI-FISH) to identify proximal (PTEC) and distal tubular epithelial cells (DTEC), whereas senescence-associated β-galactosidase (SABG) staining was used to evaluate senescence. Results revealed statistically significant decreases in the average telomere fluorescence intensity (TFI) of PTEC in CKD cats compared with young and geriatric normal cats, and in the DTEC of CKD cats compared with young normal cats. When histograms of individual TFI were compared, statistically significant decreases in the PTEC and DTEC of CKD cats were observed compared with young and geriatric normal cats. Concomitantly, a statistically significant increase in SABG staining was seen in CKD kidney samples compared with young normal cats. CKD cats tended to have increased SABG staining in the kidney compared with normal geriatric cats, but this did not reach statistical significance. No significant telomere shortening in liver or skin from any group was observed. Real-time quantitative telomeric repeat amplification protocol assessment of renal telomerase activity revealed comparable low levels of telomerase activity in all groups. Our results suggest that shortened telomeres and increased senescence in the kidneys of CKD cats may represent novel targets for interventional therapy.

The RON receptor tyrosine kinase is a member of the MET proto-oncogene family and is important for cell proliferation, differentiation, and cancer development. Here, we created a series of Madin-Darby canine kidney (MDCK) epithelial cell clones that express different levels of RON, and have investigated their biological properties. While low levels of RON correlated with little morphological change in MDCK cells, high levels of RON expression constitutively led to morphological scattering or complete and stabilized epithelial-to-mesenchymal transition (EMT). Unexpectedly, MDCK clones expressing higher levels of RON exhibited retarded proliferation and senescence, despite increased motility and invasiveness. RON was constitutively tyrosine-phosphorylated in MDCK cells expressing high levels of RON and undergoing EMT, and the MAPK signaling pathway was activated. This study reveals for the first time that RON alone is sufficient to induce complete and stabilized EMT in MDCK cells, and overexpression of RON does not cause cell transformation but rather induces cell cycle arrest and senescence, leading to impaired cell proliferation.

Nanotechnology, as a new enabling technology, has the potential to revolutionize food systems. However, much attention has been focused on nanoparticle foods due to their potential physiological properties. This study was aimed to evaluate the mutagenic safety and fatty liver improvement of black soybean in senescence-accelerated mice (SAMP8). The mutagenic activity of black soybeans was investigated using the Ames test (Salmonella Typhimurium TA98, 100, 102, and 1535). Furthermore, senescence-accelerated prone-8 mice (SAMP8) have been reported to display spontaneous fatty liver. Male SAMP8 mice were divided into control and supplemented with 10% micronized or nanonized black soybeans diet and fed for 12 wk. The results revealed that the Ames test of micronized and nanonized black soybeans exhibited no mutagenicity. Administration of black soybeans to mice showed no effects on food intake and body and organ weights. The nanonized black soybean group had a lower degree of spontaneous fatty liver, alanine aminotransferase, and thiobarbituric acid-reactive substance concentrations, and had enhanced superoxide dismutase, catalase, and glutathione peroxidase activities of livers when compared with the SAMP8 control and micronized black soybean groups. The mice fed with black soybeans had significantly lower triglyceride concentrations than the SAMP8 control group. The results of this study suggest that nanonized black soybeans have no side effects and, moreover, may minimize liver lesions in SAMP8 mice.

Sarcopenia is a condition of the loss of muscle mass that is associated with aging and that increases the risk for bedridden state, thereby warranting studies of interventions that attenuate sarcopenia. Here the effects of 2-month dietary L-lysine (Lys) supplementation (1.5-3.0 %) on myofibrillar protein degradation and major proteolytic systems were investigated in senescence-accelerated mouse prone 8 (SAMP8). At 36 weeks of age, skeletal muscle and lean body mass was reduced in SAMP8 when compared with control senescence-accelerated mouse resistant 1 (SAMR1). The myofibrillar protein degradation, which was evaluated by the release of 3-methylhistidine, was stimulated in SAMP8, and the autophagy activity, which was evaluated by light chain 3-II, was stimulated in the skeletal muscle of SAMP8. The activation of ubiquitin-proteasome system was not observed in the muscles of SAMP8. However, myofibrillar protein degradation and autophagic activity in skeletal muscles of SAMP8 were suppressed by dietary intake of 3.0 % Lys. The present data indicate that myofibrillar protein degradation by bulk autophagy is stimulated in the skeletal muscles of SAMP8 and that dietary Lys supplementation attenuates sarcopenia in SAMP8 by suppressing autophagic proteolysis.

Spirulina has proven to be effective in treating certain cancers, hyperlipidemia, immunodeficiency, and inflammatory processes. In this study, we aimed to investigate the effects of Spirulina on memory dysfunction, oxidative stress damage and antioxidant enzyme activity. Three-month-old male senescence-accelerated prone-8 (SAMP8) mice were randomly assigned to either a control group or to one of two experimental groups (one receiving daily dietary supplementation with 50 mg/kg BW and one with 200 mg/kg BW of Spirulina platensis water extract). Senescence-accelerated-resistant (SAMR1) mice were used as the external control. Results showed that the Spirulina-treated groups had better passive and avoidance scores than the control group. The amyloid β-protein (Aβ) deposition was significantly reduced at the hippocampus and whole brain in both Spirulina groups. The levels of lipid peroxidation were significantly reduced at the hippocampus, striatum, and cortex in both Spirulina groups, while catalase activity was significantly higher only in the 200 mg/kg BW Spirulina group than in the control group. Glutathione peroxidase activity was significantly higher only in the cortex of the 200 mg/kg group than in that of the SAMP8 control group. However, superoxide dismutase activity in all parts of the brain did not significantly differ among all groups. In conclusion, Spirulina platensis may prevent the loss of memory possibly by lessening Aβ protein accumulation, reducing oxidative damage and mainly augmenting the catalase activity.

Computerized morphometry was used to examine the sizes of neuronal bodies and the compactness of arrangement of neurons and neuroglial cells in layers III and V of the sensorimotor cortex in senescence-accelerated prone 1 (SAMP1) mice (an experimental group) and senescence-accelerated-resistant strain 1 (SAMR1) ones (a control group). In the SAMP1 mice as compared to the SAMR1 ones, the neuronal body sizes were significantly unchanged; the compactness of their arrangement decreased by 17 and 20% in layers III and V, respectively; that of neuroglial cells significantly increased by 14% in layer III only. In the SAMP1 mice versus the SAMR1 ones, the glial index rose by 36% in layer III and by 24% in layer V. During simulation of physiological aging, the sizes of neuronal bodies were shown to be virtually unchanged in the cerebral cortex; the compactness of their arrangement (cell counts) moderately reduced and that of neuroglial cells increased, which caused a rise in the glioneuronal index that was indicative of the enhanced supporting function of neuroglial cells during the physiological aging of brain structures.

Cardiac senescence and age-related disease development have gained general attention and recognition in the past decades due to increased accessibility and quality of health care. The advancement in global civilization is complementary to concerns regarding population aging and development of chronic degenerative diseases. Cardiac degeneration has been rigorously studied. The molecular mechanisms of cardiac senescence are on multiple cellular levels and hold a multilayer complexity level, thereby hampering development of unambiguous treatment protocols. In particular, the synergistic exchange of the senescence phenotype through a senescence secretome between myocytes and stem cells appears complicated and is of great future therapeutic value. The current review article will highlight hallmarks of senescence, cardiac myocyte and stem cell senescence, and the mutual exchange of senescent secretome. Future cardiac cell therapy approaches require a comprehensive understanding of myocardial senescence to improve therapeutic efficiency as well as efficacy. PMID:24349878

Cellularsenescence is a tumor-suppressive program that involves chromatin reorganization and specific changes in gene expression that trigger an irreversible cell-cycle arrest. Here we combine quantitative mass spectrometry, ChIP deep-sequencing, and functional studies to determine the role of histone modifications on chromatin structure and gene-expression alterations associated with senescence in primary human cells. We uncover distinct senescence-associated changes in histone-modification patterns consistent with a repressive chromatin environment and link the establishment of one of these patterns--loss of H3K4 methylation--to the retinoblastoma tumor suppressor and the H3K4 demethylases Jarid1a and Jarid1b. Our results show that Jarid1a/b-mediated H3K4 demethylation contributes to silencing of retinoblastoma target genes in senescent cells, suggesting a mechanism by which retinoblastoma triggers gene silencing. Therefore, we link the Jarid1a and Jarid1b demethylases to a tumor-suppressor network controlling cellularsenescence.

Cellularsenescence is a tumor-suppressive program that involves chromatin reorganization and specific changes in gene expression that trigger an irreversible cell-cycle arrest. Here we combine quantitative mass spectrometry, ChIP deep-sequencing, and functional studies to determine the role of histone modifications on chromatin structure and gene-expression alterations associated with senescence in primary human cells. We uncover distinct senescence-associated changes in histone-modification patterns consistent with a repressive chromatin environment and link the establishment of one of these patterns—loss of H3K4 methylation—to the retinoblastoma tumor suppressor and the H3K4 demethylases Jarid1a and Jarid1b. Our results show that Jarid1a/b-mediated H3K4 demethylation contributes to silencing of retinoblastoma target genes in senescent cells, suggesting a mechanism by which retinoblastoma triggers gene silencing. Therefore, we link the Jarid1a and Jarid1b demethylases to a tumor-suppressor network controlling cellularsenescence. PMID:22615382

Increased levels of oxidized proteins with aging have been considered a cardiovascular risk factor. However, it is unclear whether oxidized albumin, which is the most abundant serum protein, induces endothelial damage. The results of this study indicated that with aging processes, the levels of oxidized proteins as well as endothelial microparticles release increased, a novel marker of endothelial damage. Among these, oxidized albumin seems to play a principal role. Through in vitro studies, endothelial cells cultured with oxidized albumin exhibited an increment of endothelial damage markers such as adhesion molecules and apoptosis levels. In addition, albumin oxidation increased the amount of endothelial microparticles that were released. Moreover, endothelial cells with increased oxidative stress undergo senescence. In addition, endothelial cells cultured with oxidized albumin shown a reduction in endothelial cell migration measured by wound healing. As a result, we provide the first evidence that oxidized albumin induces endothelial injury which then contributes to the increase of cardiovascular disease in the elderly subjects. PMID:27042026

Photobiomodulation (PBM) is an effect oflow intensity monochromatic light or laser irradiation (LIL) on biological systems. which stimulates or inhibits biological functions but does not result in irreducible damage. It has been observed that PBM can suppress cellularsenescence, reverse skin photoageing and improve fibromyalgia. In this paper, the biological information model of photobiomodulation (BIMP) is used to discuss its mechanism. Cellularsenescence can result from short, dysfunctional telomeres, oxidative stress, or oncogene expression, and may contribute to aging so that it can be seen as a decline of cellular function in which cAMP plays an important role, which provide a foundation for PBM on senescence since cellularsenescence is a reasonable model of senescence and PBM is a cellular rehabilitation in which cAMP also plays an important role according to BIMP. The PBM in reversing skin photoageing and improving fibromyalgia are then discussed in detail.

One of the features of cellularsenescence is the activity of senescence-associated- β-galactosidase (SA-β-gal). The main purpose of this study was to evaluate this marker of senescence in aging neurons. We found that cortical neurons exhibited noticeable SA-β-gal activity quite early in culture. Many SA-β-gal-positive neurons were negative for another canonical marker of senescence, namely, double-strand DNA breaks (DSBs). Moreover, DDR signalling triggered by low doses of doxorubicin did not accelerate the appearance of neuronal SA-β-gal. In vivo, we observed pronounced induction of SA-β-gal activity in the hippocampus of 24-month-old mice, which is consistent with previous findings and supports the view that at this advanced age neurons developed a senescence-like phenotype. Surprisingly however, relatively high SA-β-gal activity, probably unrelated to the senescence process, was also observed in much younger, 3-month-old mice. In conclusion, we propose that SA-β-gal activity in neurons cannot be attributed uniquely to cell senescence either in vitro or in vivo. Additionally, we showed induction of REST protein in aging neurons in long-term culture and we propose that REST could be a marker of neuronal senescence in vitro. PMID:27768595

It has been reported that the induction of cellularsenescence through p53 activation is an effective strategy in tumor regression. Unfortunately, however, tumors including adult T-cell leukemia/lymphoma (ATL) have disadvantages such as p53 mutations and a lack of p16(INK4a) and/or p14(ARF). In this study we characterized Nutlin-3a-induced cell death in 16 leukemia/lymphoma cell lines. Eight cell lines, including six ATL-related cell lines, had wild-type p53 and Nutlin-3a-activated p53, and the cell lines underwent apoptosis or cell-cycle arrest, whereas eight cell lines with mutated p53 were resistant. Interestingly, senescence-associated-beta-galactosidase (SA-beta-gal) staining revealed that only ATL-related cell lines with wild-type p53 showed cellularsenescence, although they lack both p16(INK4a) and p14(ARF). These results indicate that cellularsenescence is an important event in p53-dependent cell death in ATL cells and is inducible without p16(INK4a) and p14(ARF). Furthermore, knockdown of Tp53-induced glycolysis and apoptosis regulator (TIGAR), a novel target gene of p53, by small interfering RNA(siRNA) indicated its important role in the induction of cellularsenescence. As many patients with ATL carry wild-type p53, our study suggests that p53 activation by Nutlin-3a is a promising strategy in ATL. We also found synergism with a combination of Nutlin-3a and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), suggesting the application of Nutlin-3a-based therapy to be broader than expected.

8-Hydroxy-2'-deoxyguanosine (8-OHdG), one of the most abundant oxidative DNA adducts, is used as an indicator of oxidative DNA damage associated with aging. Among homologs of the silent information regulator (Sir), sirtuin 1 (SIRT1) is suggested as a regulator of the apoptotic response to DNA damage. Since it has been suggested that the aging process can be delayed by the attenuation of oxidative damage such as DNA damage or SIRT1 modulation, we focused on the protective effect against cellular oxidative damage of persimmon peel, a proanthocyanidin-rich food, in relation to its level of polymerization. We confirmed that 8-OHdG expression in TIG-1 human fibroblasts was increased by treatment with 300 microM H2O2 for 2 h. On the other hand, the nuclear SIRT1 level was decreased in H2O2-treated as compared with non-pretreated cells. However, pretreatments with polymers and oligomers led to a decrease in 8-OHdG and elevation in nuclear SIRT1 expression in a concentration-dependent manner. In particular, oligomers exerted a stronger effect. The present study supports the protective potential of proanthocyanidin from persimmon peel against oxidative damage under the aging process, and suggests that the polymerization of proanthocyanidin plays an important role in retarding aging in a cellularsenescence model.

SUMMARY Cellularsenescence suppresses cancer by halting the growth of premalignant cells, yet the accumulation of senescent cells is thought to drive age-related pathology through a senescence-associated secretory phenotype (SASP), the function of which is unclear. To understand the physiological role(s) of the complex senescent phenotype, we generated a mouse model in which senescent cells can be visualized and eliminated in living animals. We show that senescent fibroblasts and endothelial cells appear very early in response to a cutaneous wound, where they accelerate wound closure by inducing myofibroblast differentiation through the secretion of platelet-derived growth factor AA (PDGF-AA). In two mouse models, topical treatment of senescence-free wounds with recombinant PDGF-AA rescued the delayed wound closure and lack of myofibroblast differentiation. These findings define a beneficial role for the SASP in tissue repair and help to explain why the SASP evolved. PMID:25499914

Cellularsenescence suppresses cancer by halting the growth of premalignant cells, yet the accumulation of senescent cells is thought to drive age-related pathology through a senescence-associated secretory phenotype (SASP), the function of which is unclear. To understand the physiological role(s) of the complex senescent phenotype, we generated a mouse model in which senescent cells can be visualized and eliminated in living animals. We show that senescent fibroblasts and endothelial cells appear very early in response to a cutaneous wound, where they accelerate wound closure by inducing myofibroblast differentiation through the secretion of platelet-derived growth factor AA (PDGF-AA). In two mouse models, topical treatment of senescence-free wounds with recombinant PDGF-AA rescued the delayed wound closure and lack of myofibroblast differentiation. These findings define a beneficial role for the SASP in tissue repair and help to explain why the SASP evolved.

Fusarium oxysporum f.sp. cubense (FOC) is a causal agent of vascular wilt and leaf chlorosis of banana plants. Chloroses resulting from FOC occur first in the lowest leaves of banana seedlings and gradually progress upward. To investigate the responses of different leaf positions to FOC infection, hydroponic experiments with FOC inoculation were conducted in a greenhouse. Fusarium-infected seedlings exhibited a decrease in net photosynthesis rate, stomatal conductance, and transpiration rate of all leaves. The wilting process in Fusarium-infected seedlings varied with leaf position. Measurements of the maximum photochemical efficiency of photosystem II (F(V)/F(max) and visualization with transmission electron microscopy showed a positive correlation between chloroplast impairment and severity of disease symptoms. Furthermore, results of malondialdehyde content and relative membrane conductivity measurements demonstrated that the membrane system was damaged in infected leaves. Additionally, the activities of phenylalanine ammonia-lyase, peroxidase and polyphenol oxidase were increased and total soluble phenolic compounds were significantly accumulated in the leaves of infected plants. The structural and biochemical changes of infected plants was consistent with plant senescence. As the FOC was not detected in infected leaves, we proposed that the chloroplast and membrane could be damaged by fusaric acid produced by Fusarium. During the infection, fusaric acid was first accumulated in the lower leaves and water-soluble substances in the lower leaves could dramatically enhance fusaric acid production. Taken together, the senescence of infected banana plants was induced by Fusarium infection with fusaric acid production and the composition of different leaf positions largely contribute to the particular senescence process.

In this study, we tested the hypotheses that endothelial cells (ECs) derived from human umbilical cord blood (hCB-ECs) exhibit low permeability, which increases as hCB-ECs age and undergo senescence, and that the change in the permeability of hCB-ECs is due to changes in tight junction protein localization and the activity of exchange protein activated by cAMP (Epac)1. Albumin permeability across low-passage hCB-EC monolayers on Transwell membranes was 10 times lower than for human aortic ECs (HAECs) (P < 0.01) but similar to in vivo values in arteries. Expression of the tight junction protein occludin and tyrosine phosphorylation of occludin were less in hCB-ECs than in HAECs (P < 0.05). More hCB-ECs than HAECs underwent mitosis (P < 0.01). hCB-ECs that underwent >44 population doublings since isolation had a significantly higher permeability than hCB-ECs that underwent <31 population doublings (P < 0.05). This age-related increase in hCB-EC permeability was associated with an increase in tyrosine phosphorylation of occludin (P < 0.01); permeability and occludin phosphorylation were reduced by treatment with 2 μM resveratrol. Tyrosine phosphorylation of occludin and cell age influence the permeability of hCB-ECs, whereas levels of EC proliferation and expression of tight junction proteins did not explain the differences between hCB-EC and HAEC permeability. The elevated permeability in late passage hCB-ECs was reduced by 25-40% by elevation of membrane-associated cAMP and activation of the Epac1 pathway. Given the similarity to in vivo permeability to albumin and the high proliferation potential, hCB-ECs may be a suitable in vitro model to study transport-related pathologies and cell aging.

This study has investigated the effect of aging on parameters of inflammation, oxidative stress and apoptosis in pancreas obtained from two types of male mice models: senescence-accelerated prone (SAMP8) and resistant mice (SAMR1). Animals of 2 (young) and 10 months of age (old) were used (n = 64). The influence of the administration of melatonin in the drinking water for one month at two different dosages (1 and 10mg/(kg day) on old SAMP8 mice on these parameters was also studied. SAMP8 mice showed with age a significant increase in the relative expression of pancreatic genes involved in inflammation, oxidative stress and apoptosis. Furthermore the protein expression of several NFκB subunits was also enhanced. On the contrary aged SAMR1 mice did not show significant increases in these parameters. Melatonin administration to SAMP8 mice was able to reduce these age related alterations at the two used dosages.

Senescence is an irreversible proliferation arrest that is induced by various stress stimuli including genotoxin. Pneumolysin (PLY) is a pathogenicity factor unique to Streptococcus pneumoniae that is important in pneumococcal-induced diseases such as otitis media, meningitis and pneumonia. However, the cell fate response to the toxin is mechanistically unclear. We investigated the effect of PLY on cellularsenescence in BV-2 microglial cells. Exposure to PLY resulted in changes in the expression of phospho-p53, p21, p16, pRb and CDK2 and increased the number of senescence associated β-gal positive cells. PLY-treatment also increased PAI-1 expression and cell proliferation arrest in concentration- and time-dependent manners. PLY induced NF-κB activation and phosphorylation of SIRT-1, ERK1/2, JNK, and p38 MAPK. In addition, PLY increased the production of reactive oxygen species. Overall, the results suggest that PLY regulates microglial cellularsenescence by enhancing production of reactive oxygen species, activation of MAPK and NF-κB, and phosphorylation of SIRT-1.

The aim of the present study was to investigate the effect of aging on several parameters related to glucose homeostasis and insulin resistance in pancreas and how melatonin administration could affect these parameters. Pancreas samples were obtained from two types of male mice models: senescence-accelerated prone (SAMP8) and senescence-accelerated-resistant mice (SAMR1). Insulin levels in plasma were increased with aging in both SAMP8 and SAMR1 mice, whereas insulin content in pancreas was decreased with aging in SAMP8 and increased in SAMR1 mice. Expressions of glucagon and GLUT2 messenger RNAs (mRNAs) were increased with aging in SAMP8 mice, and no differences were observed in somatostatin and insulin mRNA expressions. Furthermore, aging decreased also the expressions of Pdx-1, FoxO 1, FoxO 3A and Sirt1 in pancreatic SAMP8 samples. Pdx-1 was decreased in SAMR1 mice, but no differences were observed in the rest of parameters on these mice strains. Treatment with melatonin was able to decrease plasma insulin levels and to increase its pancreatic content in SAMP8 mice. In SAMR1, insulin pancreatic content and plasma levels were decreased. HOMA-IR was decreased with melatonin treatment in both strains of animals. On the other hand, in SAMP8 mice, treatment decreased the expression of glucagon, GLUT2, somatostatin and insulin mRNA. Furthermore, it was also able to increase the expression of Sirt1, Pdx-1 and FoxO 3A. According to these results, aging is associated with significant alterations in the relative expression of pancreatic genes associated to glucose metabolism. This has been especially observed in SAMP8 mice. Melatonin administration was able to improve pancreatic function in old SAMP8 mice and to reduce HOMA-IR improving their insulin physiology and glucose metabolism.

Cellularsenescence is an important mechanism for preventing tumor progression. The elevated expression of Bcl-2-interacting cell death suppressor (BIS), an anti-apoptotic and anti-stress protein, often correlates with poor prognosis in several cancers including glioblastoma; however, the role of BIS in the regulation of senescence has not been well defined. Here, we describe for the first time that the depletion of BIS induces G1 arrest and cellularsenescence through the accumulation of p27 that is independent of p53, p21 or p16. The increase in p27 expression in BIS-depleted cells was attributable to an impairment of the ubiquitin-mediated degradation of p27, which was caused by a decrease in S-phase kinase-associated protein 2 (SKP2) at the transcriptional level. As an underlying molecular mechanism, we demonstrate that the loss of activity of signal transducer and activator of transcription 3 (STAT3) was specifically linked to the suppression of SKP2 expression. Despite a reduction in phospho-STAT3 levels, total STAT3 levels were unexpectedly increased by BIS depletion, specifically in the insoluble fraction. Our results show that 14-3-3ζ expression is decreased by BIS knockdown and that 14-3-3ζ depletion per se significantly induced senescence phenotypes. In addition, the ectopic expression of 14-3-3ζ blocked senescence caused by BIS depletion, which was paralleled with a decrease in insoluble STAT3 in A172 glioblastoma cells. These findings indicate that the impairment of the protein quality control conferred by BIS and/or 14-3-3ζ is critical for BIS depletion-induced senescence. Moreover, BIS knockdown also induced senescence along with an accumulation of total STAT3 and p27 in several different cell types as well as embryonic fibroblasts derived from Bis-knock out mice with/without variations in 14-3-3ζ levels. Therefore, our findings suggest that a downregulation of BIS expression could serve as a potential strategy for restricting tumor progression

Activation of NOTCH signalling is associated with advanced prostate cancer and treatment resistance in prostate cancer patients. However, the mechanism that drives NOTCH activation in prostate cancer remains still elusive. Moreover, preclinical evidence of the therapeutic efficacy of NOTCH inhibitors in prostate cancer is lacking. Here, we provide evidence that PTEN loss in prostate tumours upregulates the expression of ADAM17, thereby activating NOTCH signalling. Using prostate conditional inactivation of both Pten and Notch1 along with preclinical trials carried out in Pten-null prostate conditional mouse models, we demonstrate that Pten-deficient prostate tumours are addicted to the NOTCH signalling. Importantly, we find that pharmacological inhibition of γ-secretase promotes growth arrest in both Pten-null and Pten/Trp53-null prostate tumours by triggering cellularsenescence. Altogether, our findings describe a novel pro-tumorigenic network that links PTEN loss to ADAM17 and NOTCH signalling, thus providing the rational for the use of γ-secretase inhibitors in advanced prostate cancer patients. PMID:27941799

The influence of two known cardiovascular risk factors, aging and consumption of a high-fat diet, on vascular mesenteric artery reactivity was examined in a mouse model of acceleratedsenescence (SAM). Five-month-old SAM prone (SAMP8) and resistant (SAMR1) female mice were fed a Western-type high-fat diet (WD; 8 weeks). Mesenteric arteries were dissected, and vascular reactivity, protein and messenger RNA expression, superoxide anion (O 2 (·-) ) and hydrogen peroxide formation were evaluated by wire myography, immunofluorescence, RT-qPCR, ethidium fluorescence and ferric-xylenol orange, respectively. Contraction to KCl and relaxation to acetylcholine remained unchanged irrespective of senescence and diet. Although similar contractions to phenylephrine were observed in SAMR1 and SAMP8, acceleratedsenescence was associated with decreased eNOS and nNOS and increased O 2 (·-) synthesis. Senescence-related alterations were compensated, at least partly, by the contribution of NO derived from iNOS and the enhanced endogenous antioxidant capacity of superoxide dismutase 1 to maintain vasoconstriction. Administration of a WD induced qualitatively different alterations in phenylephrine contractions of mesenteric arteries from SAMR1 and SAMP8. SAMR1 showed increased contractions partly as a result of decreased NO availability generated by decreased eNOS and nNOS and enhanced O 2 (·-) formation. In contrast, WD feeding in SAMP8 resulted in reduced contractions due to, at least in part, the increased functional participation of iNOS-derived NO. In conclusion, senescence-dependent intrinsic alterations during early stages of vascular senescence may promote vascular adaptation and predispose to further changes in response to high-fat intake, which may lead to the progression of aging-related cardiovascular disease, whereas young subjects lack the capacity for this adaptation.

The risk of cardiovascular complications in diabetic patients is mainly associated with endothelial dysfunction. Reduced number of EPCs and impaired function of EPCs in diabetes result in imbalance of endothelial homeostasis and dysfunction of vessels. In patients with diabetes mellitus, plasma levels of asymmetric dimethylarginine (ADMA) were elevated, while the expression and activity of dimethylarginine dimethylaminohydrolase (DDAH) were reduced. In the present study, we investigated the role of the DDAH2/ADMA pathway in the senescence of EPCs in type 2 diabetic patients and cultured EPCs treated with high glucose. The results showed that the percentage of senescent EPCs increased while the expression of DDAH2 decreased concomitantly with an increase in the plasma levels of ADMA in patients with type 2 diabetes mellitus (T2DM). Similar results were seen in cultured EPCs treated with high glucose. Exogenous application of ADMA accelerated the senescence of EPCs in a dose-dependent manner, and overexpression of DDAH2 inhibited high glucose-induced EPCs senescence. In addition, it has also been reported that DDAH/ADMA pathway is regulated by silent information regulator 1 (SIRT1) in endothelial cell. In the present study, we found decreased expression of SIRT1 both in T2DM patients and EPCs pretreated with high glucose. And resveratrol (activating SIRT1) inhibited high glucose-induced EPCs senescence by upregulating the expression of DDAH2 and decreasing the levels of ADMA. Taken together, we concluded that DDAH2/ADMA is involved in the acceleratedsenescence of EPCs in diabetes, which is associated with the activation of SIRT1.

We studied potential changes in the subventricular zone (SVZ) stem cell niche of the senescence-accelerated mouse prone-8 (SAM-P8) aging model. Bromodeoxyuridine (BrdU) assays with longtime survival revealed a lower number of label-retaining stem cells in the SAM-P8 SVZ compared with the SAM-Resistant 1 (SAM-R1) control strain. We also found that in SAM-P8 niche signaling is attenuated and the stem cell pool is less responsive to the self-renewal niche factor pigmented epithelium-derived factor (PEDF). Protein analysis demonstrated stable amounts of the PEDF ligand in the SAM-P8 SVZ niche; however, SAM-P8 stem cells present a significant expression decrease of patatin-like phospholipase domain containing 2, a receptor for PEDF (PNPLA2-PEDF) receptor, but not of laminin receptor (LR), a receptor for PEDF (LR-PEDF) receptor. We observed changes in self-renewal related genes (hairy and enhancer of split 1 (Hes1), hairy and enhancer of split 1 (Hes5), Sox2] and report that although these genes are down-regulated in SAM-P8, differentiation genes (Pax6) are up-regulated and neurogenesis is increased. Finally, sheltering mammalian telomere complexes might be also involved given a down-regulation of telomeric repeat binding factor 1 (Terf1) expression was observed in SAM-P8 at young age periods. Differences between these 2 models, SAM-P8 and SAM-R1 controls, have been previously detected at more advanced ages. We now describe alterations in the PEDF signaling pathway and stem cell self-renewal at a very young age, which could be involved in the premature senescence observed in the SAM-P8 model.

Introduction Dehydroepiandrosterone (DHEA), an adrenal 17-ketosteroid, is a precursor of testosterone and 17β-estradiol. Studies have shown that DHEA inhibits carcinogenesis in mammary gland and prostate as well as other organs, a process that is not hormone dependent. Little is known about the molecular mechanisms of DHEA-mediated inhibition of the neoplastic process. Here we examine whether DHEA and its analog DHEA 8354 can suppress the progression of hyperplastic and premalignant (carcinoma in situ) lesions in mammary gland toward malignant tumors and the cellular mechanisms involved. Methods Rats were treated with N-nitroso-N-methylurea and allowed to develop mammary hyperplastic and premalignant lesions with a maximum frequency 6 weeks after carcinogen administration. The animals were then given DHEA or DHEA 8354 in the diet at 125 or 1,000 mg/kg diet for 6 weeks. The effect of these agents on induction of apoptosis, senescence, cell proliferation, tumor burden and various effectors of cellular signaling were determined. Results Both agents induced a dose-dependent decrease in tumor multiplicity and in tumor burden. In addition they induced a senescent phenotype in tumor cells, inhibited cell proliferation and increased the number of apoptotic cells. The DHEA-induced cellular effects were associated with increased expression of p16 and p21, but not p53 expression, implicating a p53-independent mechanism in their action. Conclusion We provide evidence that DHEA and DHEA 8354 can suppress mammary carcinogenesis by altering various cellular functions, inducing cellularsenescence, in tumor cells with the potential involvement of p16 and p21 in mediating these effects. PMID:16457693

Aging is associated with an increase in oxidative stress and inflammation. The aging lung is particularly affected since it is continuously exposed to environmental oxidants while antioxidant machinery weakens with age. Melatonin, a free radical scavenger, counteracts inflammation and apoptosis in healthy cells from several tissues. Its effects on the aging lung are, however, not yet fully understood. This study aimed to investigate the effect of chronic administration of melatonin on the expression of inflammation markers (TNF-α, IL-1β, NFκB2, HO-1) and apoptosis parameters (BAD, BAX, AIF) in the lung tissue of male senescence-accelerated prone mice (SAMP8). In addition, RNA oxidative damage, as the formation of 8-hydroxyguanosine (8-OHG), was also evaluated. Young and old animals, aged 2 and 10 months respectively, were divided into 4 groups: untreated young, untreated old, old mice treated with 1mg/kg/day melatonin, and old animals treated with 10mg/kg/day melatonin. Untreated young and old male senescenceaccelerated resistant mice (SAMR1) were used as controls. After 30 days of treatment, animals were sacrificed. Lungs were collected and immediately frozen in liquid nitrogen. mRNA and protein expressions were measured by RT-PCR and Western blotting, respectively. Levels of 8-OHG were quantified by ELISA. Mean values were analyzed using ANOVA. Old nontreated SAMP8 animals showed increased (p<0.05) mRNA and protein levels of TNF-α, IL-1β, NFκB2, and HO-1 compared to young mice and SAMR1 mice. Melatonin treatment with either dose reversed the aging-derived inflammation (p<0.05). BAD, BAX and AIF expressions also rose with aging, the effect being counteracted with melatonin (p<0.05). Aging also caused a significant elevation (p<0.05) in SAMP8 8-OHG values. This increase was not observed in animals treated with melatonin (p<0.05). In conclusion, melatonin treatment was able to modulate the inflammatory and apoptosis status of the aging lungs, exerting a

Background Etiology of complex disorders, such as cataract and neurodegenerative diseases including age-related macular degeneration (AMD), remains poorly understood due to the paucity of animal models, fully replicating the human disease. Previously, two quantitative trait loci (QTLs) associated with early cataract, AMD-like retinopathy, and some behavioral aberrations in senescence-accelerated OXYS rats were uncovered on chromosome 1 in a cross between OXYS and WAG rats. To confirm the findings, we generated interval-specific congenic strains, WAG/OXYS-1.1 and WAG/OXYS-1.2, carrying OXYS-derived loci of chromosome 1 in the WAG strain. Both congenic strains displayed early cataract and retinopathy but differed clinically from OXYS rats. Here we applied a high-throughput RNA sequencing (RNA-Seq) strategy to facilitate nomination of the candidate genes and functional pathways that may be responsible for these differences and can contribute to the development of the senescence-accelerated phenotype of OXYS rats. Results First, the size and map position of QTL-derived congenic segments were determined by comparative analysis of coding single-nucleotide polymorphisms (SNPs), which were identified for OXYS, WAG, and congenic retinal RNAs after sequencing. The transferred locus was not what we expected in WAG/OXYS-1.1 rats. In rat retina, 15442 genes were expressed. Coherent sets of differentially expressed genes were identified when we compared RNA-Seq retinal profiles of 20-day-old WAG/OXYS-1.1, WAG/OXYS-1.2, and OXYS rats. The genes most different in the average expression level between the congenic strains included those generally associated with the Wnt, integrin, and TGF-β signaling pathways, widely involved in neurodegenerative processes. Several candidate genes (including Arhgap33, Cebpg, Gtf3c1, Snurf, Tnfaip3, Yme1l1, Cbs, Car9 and Fn1) were found to be either polymorphic in the congenic loci or differentially expressed between the strains. These genes may

Current understanding points to unrepairable chromosomal damage as the critical determinant of acceleratedsenescence in cancer cells treated with radiation or chemotherapy. Nonetheless, the potent senescence inducer etoposide not only targets topoisomerase II to induce DNA damage but also produces abundant free radicals, increasing cellular reactive oxygen species (ROS). Toward examining roles for DNA damage and oxidative stress in therapy-induced senescence, we developed a quantitative flow cytometric senescence assay and screened 36 redox-active agents as enhancers of an otherwise ineffective dose of radiation. While senescence failed to correlate with total ROS, the radiation enhancers, etoposide and the other effective topoisomerase inhibitors each produced high levels of lipid peroxidation. The reactive aldehyde 4-hydroxy-2-nonenal, a lipid peroxidation end product, was sufficient to induce senescence in irradiated cells. In turn, sequestering aldehydes with hydralazine blocked effects of etoposide and other senescence inducers. These results suggest that lipid peroxidation potentiates DNA damage from radiation and chemotherapy to drive therapy-induced senescence. PMID:27453792

We report herein that pulmonary emphysematous lesions appear to be a dynamic phenomenon that involves not only the gradual loss of alveolar structure but also apoptosis, cellular proliferation, and cellularsenescence. Cellular proliferation compensates for increased alveolar cell apoptosis in patients with chronic obstructive pulmonary disease (COPD). However, smoking, age, and the increased cell cycle turnover that compensates for apoptosis accelerate alveolar cell senescence, thereby halting cellular proliferation and tipping the balance toward apoptosis, which, in turn, promotes the formation of emphysematous lesions. As a result, alveolar cells disappear and the emphysematous lesions progress. At the same time, cellularsenescence is believed to induce inflammation. More specifically, senescent alveolar cells induce inflammation by producing various inflammatory cytokines in tissue. Lymphocytes and Clara cells may also age more rapidly in the lungs of patients with COPD. Lymphocyte senescence may induce an autoimmune reaction and increase susceptibility to infection, and Clara cell senescence may impair airway regeneration as well as sustain airway inflammation. Thus, cellularsenescence may be involved in arrested tissue repair, chronic inflammation, and increased susceptibility to infection, which are the typical features of COPD.

Increases in the number of patients with dementia involving Alzheimer's disease (AD) are seen as a grave public health problem. In neurodegenerative disorders involving AD, biological stresses, such as oxidative and inflammatory stress, induce neural cell damage. Asparagus (Asparagus officinalis) is a popular vegetable, and an extract prepared from this reportedly possesses various beneficial biological activities. In the present study, we investigated the effects of enzyme-treated asparagus extract (ETAS) on neuronal cells and early cognitive impairment of senescence-accelerated mouse prone 8 (SAMP8) mice. The expression of mRNAs for factors that exert cytoprotective and anti-apoptotic functions, such as heat-shock protein 70 and heme oxygenase-1, was upregulated in NG108-15 neuronal cells by treatment with ETAS. Moreover, when release of lactate dehydrogenase from damaged NG108-15 cells was increased for cells cultured in medium containing either the nitric oxide donor sodium nitroprusside or the hypoxia mimic reagent cobalt chloride, ETAS significantly attenuated this cell damage. Also, when contextual fear memory, which is considered to be a hippocampus-dependent memory, was significantly impaired in SAMP8 mice, ETAS attenuated the cognitive impairment. These results suggest that ETAS produces cytoprotective effects in neuronal cells and attenuates the effects on the cognitive impairment of SAMP8 mice.

Hybrid chlorosis, a type of hybrid incompatibility, has frequently been reported in inter- and intraspecific crosses of allopolyploid wheat. In a previous study, we reported some types of growth abnormalities such as hybrid necrosis and observed hybrid chlorosis with mild or severe abnormalities in wheat triploids obtained in crosses between tetraploid wheat cultivar Langdon and four Ae. tauschii accessions and in their derived synthetic hexaploids. However, the molecular mechanisms underlying hybrid chlorosis are not well understood. Here, we compared cytology and gene expression in leaves to characterize the abnormal growth in wheat synthetics showing mild and severe chlorosis. In addition, we compared disease resistance to wheat blast fungus. In total 55 and 105 genes related to carbohydrate metabolism and 53 and 89 genes for defense responses were markedly up-regulated in the mild and severe chlorosis lines, respectively. Abnormal chloroplasts formed in the mesophyll cells before the leaves yellowed in the hybrid chlorosis lines. The plants with mild chlorosis showed increased resistance to wheat blast and powdery mildew fungi, although significant differences only in two, third internode length and maturation time, out of the examined agricultural traits were found between the wild type and plants showing mild chlorosis. These observations suggest that senescence might be accelerated in hybrid chlorosis lines of wheat synthetics. Moreover, in wheat synthetics showing mild chlorosis, the negative effects on biomass can be minimized, and they may show substantial fitness under pathogen-polluted conditions. PMID:25806790

Oncogene-induced senescence is an anti-proliferative stress response program that acts as a fail-safe mechanism to limit oncogenic transformation and is regulated by the retinoblastoma protein (RB) and p53 tumor suppressor pathways. We identify the atypical E2F family member E2F7 as the only E2F transcription factor potently up-regulated during oncogene-induced senescence, a setting where it acts in response to p53 as a direct transcriptional target. Once induced, E2F7 binds and represses a series of E2F target genes and cooperates with RB to efficiently promote cell cycle arrest and limit oncogenic transformation. Disruption of RB triggers a further increase in E2F7, which induces a second cell cycle checkpoint that prevents unconstrained cell division despite aberrant DNA replication. Mechanistically, E2F7 compensates for the loss of RB in repressing mitotic E2F target genes. Together, our results identify a causal role for E2F7 in cellularsenescence and uncover a novel link between the RB and p53 pathways.

Oncogene-induced senescence is an anti-proliferative stress response program that acts as a fail-safe mechanism to limit oncogenic transformation and is regulated by the retinoblastoma protein (RB) and p53 tumor suppressor pathways. We identify the atypical E2F family member E2F7 as the only E2F transcription factor potently up-regulated during oncogene-induced senescence, a setting where it acts in response to p53 as a direct transcriptional target. Once induced, E2F7 binds and represses a series of E2F target genes and cooperates with RB to efficiently promote cell cycle arrest and limit oncogenic transformation. Disruption of RB triggers a further increase in E2F7, which induces a second cell cycle checkpoint that prevents unconstrained cell division despite aberrant DNA replication. Mechanistically, E2F7 compensates for the loss of RB in repressing mitotic E2F target genes. Together, our results identify a causal role for E2F7 in cellularsenescence and uncover a novel link between the RB and p53 pathways. PMID:22802529

Growth arrest is one of the essential features of cellularsenescence. At present, the precise mechanisms responsible for the establishment of the senescence-associated arrested phenotype are still incompletely understood. Given that ERK1/2 is one of the major kinases controlling cell growth and proliferation, we examined the possible implication of ERK1/2. Exposure of normal rat epithelial cells to etoposide caused cellularsenescence, as manifested by enlarged cell size, a flattened cell body, reduced cell proliferation, enhanced β-galactosidase activity, and elevated p53 and p21. Senescent cells displayed a blunted response to growth factor-induced cell proliferation, which was preceded by impaired ERK1/2 activation. Further analysis revealed that senescent cells expressed a significantly higher level of mitogen-activated protein phosphatase 3 (MKP-3, a cytosolic ERK1/2-targeted phosphatase), which was suppressed by blocking the transcriptional activity of the tumor suppressor p53 with pifithrin-α. Inhibition of MKP-3 activity with a specific inhibitor or siRNA enhanced basal ERK1/2 phosphorylation and promoted cell proliferation. Apart from its role in growth arrest, impairment of ERK1/2 also contributed to the resistance of senescent cells to oxidant-elicited cell injury. These results therefore indicate that p53-mediated up-regulation of MKP-3 contributes to the establishment of the senescentcellular phenotype through dephosphorylating ERK1/2. Impairment of ERK1/2 activation could be an important mechanism by which p53 controls cellularsenescence. PMID:25414256

Nitric oxide (NO) has been known to preserve the level of chlorophyll (Chl) during leaf senescence. However, the mechanism by which NO regulates Chl breakdown remains unknown. Here we report that NO negatively regulates the activities of Chl catabolic enzymes during dark-induced leaf senescence. The transcriptional levels of the major enzyme genes involving Chl breakdown pathway except for RED CHL CATABOLITE REDUCTASE (RCCR) were dramatically up-regulated during dark-induced Chl degradation in the leaves of Arabidopsis NO-deficient mutant nos1/noa1 that exhibited an early-senescence phenotype. The activity of pheide a oxygenase (PAO) was higher in the dark-induced senescent leaves of nos1/noa1 compared with wild type. Furthermore, the knockout of PAO in nos1/noa1 background led to pheide a accumulation in the double mutant pao1 nos1/noa1, which retained the level of Chl during dark-induced leaf senescence. The accumulated pheide a in darkened leaves of pao1 nos1/noa1 was likely to inhibit the senescence-activated transcriptional levels of Chl catabolic genes as a feed-back inhibitory effect. We also found that NO deficiency led to decrease in the stability of photosynthetic complexes in thylakoid membranes. Importantly, the accumulation of pheide a caused by PAO mutations in combination with NO deficiency had a synergistic effect on the stability loss of thylakoid membrane complexes in the double mutant pao1 nos1/noa1 during dark-induced leaf senescence. Taken together, our findings have demonstrated that NO is a novel negative regulator of Chl catabolic pathway and positively functions in maintaining the stability of thylakoid membranes during leaf senescence. PMID:23418559

A subset of AKAPs (A Kinase Anchoring Proteins) regulate signaling and cytoskeletal pathways through the spaciotemporal scaffolding of multiple protein kinases (PK), such as PKC and PKA, and associations with the plasma membrane and the actin-based cytoskeleton. SSeCKS/Gravin/Akap12 expression is severely downregulated in many advanced cancers and exhibits tumor- and metastasis-suppressing activity. akap12-null (KO) mice develop prostatic hyperplasia with focal dysplasia, but the precise mechanism how Akap12 prevents oncogenic progression remains unclear. Here, we show that KO mouse embryonic fibroblasts (MEF) exhibit premature senescence marked by polyploidy and multinucleation, and by increased susceptibility to oncogenic transformation. Although p53 and Rb pathways are activated in the absence of Akap12, senescence is dependent on Rb. Senescence is driven by the activation of PKCα, which induces p16Ink4a/Rb through a MEK-dependent downregulation of Id1, and PKCδ, which downregulates Lats1/Warts, a mitotic exit network kinase required for cytokinesis. Our data strongly suggest that Akap12 controls Rb-mediated cell aging and oncogenic progression by directly scaffolding and attenuating PKCα/δ. PMID:21099353

-related genes allow a framework to be constructed of decisive events in the evolution of the senescence syndrome of modern land-plants. Combining phylogenetic, comparative sequence, gene expression and morphogenetic information leads to the conclusion that biochemical, cellular, integrative and adaptive systems were progressively added to the ancient primary core process of senescence as the evolving plant encountered new environmental and developmental contexts. PMID:19602260

H-Ras oncogene requires deregulation of additional oncogenes or inactivation of tumor suppressor proteins to increase cell proliferation rate and transform cells. In fact, the expression of the constitutively activated H-RasV12 induces cell growth arrest and premature senescence, which act like barriers in pre-neoplastic lesions. In our experimental model, human fibroblasts transfected with H-RasV12 show a dramatic modification of morphology. H-RasV12 expressing cells also show premature senescence followed by cell death, induced by autophagy and apoptosis. In this context, we provide evidence that in H-RasV12 expressing cells, the premature senescence is associated with cellular redox imbalance as well as with altered post-translation protein modification. In particular, redox imbalance is due to a strong reduction of total antioxidant capacity, and significant decrease of glutathione level. As the reversible addition of glutathione to cysteinyl residues of proteins is an important post-translational regulative modification, we investigated S-glutathionylation in cells expressing active H-Ras. In this contest we observed different S-glutathionylation patterns in control and H-RasV12 expressing cells. Particularly, the GAPDH enzyme showed S-glutathionylation increase and significant enzyme activity depletion in H-Ras V12 cells. In conclusion, we proposed that antioxidant defense reduction, glutathione depletion and subsequent modification of S-glutathionylation of target proteins contribute to arrest cell growth, leading to death of fibroblasts expressing constitutively active H-Ras oncogene, thus acting as oncogenic barriers that obstacle the progression of cell transformation. PMID:23284910

Aging is related to loss of functional stem cell accompanying loss of tissue and organ regeneration potentials. Previously, we demonstrated that the life span of ovariectomy-senescenceaccelerated mice (OVX-SAMP8) was significantly prolonged and similar to that of the congenic senescence-resistant strain of mice after platelet rich plasma (PRP)/embryonic fibroblast transplantation. The aim of this study is to investigate the potential of PRP for recovering cellular potential from senescence and then delaying animal aging. We first examined whether stem cells would be senescent in aged mice compared to young mice. Primary adipose derived stem cells (ADSCs) and bone marrow derived stem cells (BMSCs) were harvested from young and aged mice, and found that cell senescence was strongly correlated to animal aging. Subsequently, we demonstrated that PRP could recover cell potential from senescence, such as promote cell growth (cell proliferation and colony formation), increase osteogenesis, decrease adipogenesis, restore cell senescence related markers and resist the oxidative stress in stem cells from aged mice. The results also showed that PRP treatment in aged mice could delay mice aging as indicated by survival, body weight and aging phenotypes (behavior and gross morphology) in term of recovering the cellular potential of their stem cells compared to the results on aged control mice. In conclusion these findings showed that PRP has potential to delay aging through the recovery of stem cell senescence and could be used as an alternative medicine for tissue regeneration and future rejuvenation.

Mesenchymal stromal cells (MSCs) are multipotent progenitor cells and there is much interest in how MSCs contribute to the regulation of the tumor microenvironment. Whether MSCs exert a supportive or suppressive effect on tumor progression is still controversial, but is likely dependent on a variety of factors that are tumor-type dependent. Multiple myeloma (MM) is characterized by growth of malignant plasma cells in the bone marrow. It has been shown that the progression of MM is governed by MSCs, which act as a stroma of the myeloma cells. Although stroma is created via mutual communication between myeloma cells and MSCs, the mechanism is poorly understood. Here we explored the role of lysophosphatidic acid (LPA) signaling in cellular events where MSCs were converted into either MM-supportive or MM-suppressive stroma. We found that myeloma cells stimulate MSCs to produce autotaxin, an indispensable enzyme for the biosynthesis of LPA, and LPA receptor 1 (LPA1) and 3 (LPA3) transduce opposite signals to MSCs to determine the fate of MSCs. LPA3-silenced MSCs (siLPA3-MSCs) exhibited cellularsenescence-related phenotypes in vitro, and significantly promoted progression of MM and tumor-related angiogenesis in vivo. In contrast, siLPA1-MSCs showed resistance to cellularsenescence in vitro, and efficiently delayed progression of MM and tumor-related angiogenesis in vivo. Consistently, anti-MM effects obtained by LPA1-silencing in MSCs were completely reproduced by systemic administration of Ki6425, an LPA1 antagonist. Collectively, our results indicate that LPA signaling determines the fate of MSCs and has potential as a therapeutic target in MM. Stem Cells 2017;35:739-753.

Progressively sophisticated understanding of cellular and molecular processes that contribute to age-related physical deterioration is being gained from ongoing research into cancer, chronic inflammatory syndromes and other serious disorders that increase with age. Particularly valuable insight has resulted from characterization of how senescent cells affect the tissues in which they form in ways that decrease an organism's overall viability. Increasingly, the underlying pathophysiology of ageing is recognized as a consequence of oxidative damage. This leads to hyperactivity of cell growth pathways, prominently including mTOR (mammalian target of rapamycin), that contribute to a build-up in cells of toxic aggregates such as progerin (a mutant nuclear cytoskeletal protein), lipofuscin and other cellular debris, triggering formation of senescentcellular phenotypes, which interact destructively with surrounding tissue. Indeed, senescent cell ablation dramatically inhibits physical deterioration in progeroid (age-accelerated) mice. This review explores ways in which oxidative stress creates ageing-associated cellular damage and triggers induction of the cell death/survival programs’ apoptosis, necrosis, autophagy and ‘necroapoptophagy’. The concept of ‘necroapoptophagy’ is presented here as a strategy for varying tissue oxidative stress intensity in ways that induce differential activation of death versus survival programs, resulting in enhanced and sustained representation of healthy functional cells. These strategies are discussed in the context of specialized mesenchymal stromal cells with the potential to synergize with telocytes in stabilizing engrafted progenitor cells, thereby extending periods of healthy life. Information and concepts are summarized in a hypothetical approach to suppressing whole-organism senescence, with methods drawn from emerging understandings of ageing, gained from Cnidarians (jellyfish, corals and anemones) that undergo a

Studying the phenomenon of cellularsenescence has been hindered by the lack of senescence-specific markers. As such, detection of proteins informally associated with senescence accompanies the use of senescence-associated β-galactosidase as a collection of semiselective markers to monitor the presence of senescent cells. To identify novel biomarkers of senescence, we immunized BALB/c mice with senescent mouse lung fibroblasts and screened for antibodies that recognized senescence-associated cell-surface antigens by FACS analysis and a newly developed cell-based ELISA. The majority of antibodies that we isolated, cloned, and sequenced belonged to the IgM isotype of the innate immune system. In-depth characterization of one of these monoclonal, polyreactive natural antibodies, the IgM clone 9H4, revealed its ability to recognize the intermediate filament vimentin. By using 9H4, we observed that senescent primary human fibroblasts express vimentin on their cell surface, and MS analysis revealed a posttranslational modification on cysteine 328 (C328) by the oxidative adduct malondialdehyde (MDA). Moreover, elevated levels of secreted MDA-modified vimentin were detected in the plasma of aged senescence-accelerated mouse prone 8 mice, which are known to have deregulated reactive oxygen species metabolism and accelerated aging. Based on these findings, we hypothesize that humoral innate immunity may recognize senescent cells by the presence of membrane-bound MDA-vimentin, presumably as part of a senescence eradication mechanism that may become impaired with age and result in senescent cell accumulation. PMID:28193858

The emergence of resistance to imatinib mediated by mutations in the BCR-ABL has become a major challenge in the treatment of chronic myeloid leukemia (CML). Alternative therapeutic strategies to override imatinib-resistant CML are urgently needed. In this study, we investigated the effect of AKI603, a novel small molecule inhibitor of Aurora kinase A (AurA) to overcome resistance mediated by BCR-ABL-T315I mutation. Our results showed that AKI603 exhibited strong anti-proliferative activity in leukemic cells. AKI603 inhibited cell proliferation and colony formation capacities in imatinib-resistant CML cells by inducing cell cycle arrest with polyploidy accumulation. Surprisingly, inhibition of AurA by AKI603 induced leukemia cell senescence in both BCR-ABL wild type and T315I mutation cells. Furthermore, the induction of senescence was associated with enhancing reactive oxygen species (ROS) level. Moreover, the anti-tumor effect of AKI603 was proved in the BALB/c nude mice KBM5-T315I xenograft model. Taken together, our data demonstrate that the small molecule AurA inhibitor AKI603 may be used to overcome drug resistance induced by BCR-ABL-T315I mutation in CML. PMID:27824120

To enhance the effect of anti-influenza-virus agent treatment, the effect of combined administration of oseltamivir phosphate and hochu-ekki-to (Japanese traditional herbal medicine, HET) on early viral clearance was examined. Senescence-accelerated mice were given HET in drinking water for 2 weeks, followed by intranasal infection with influenza A virus strain PR8. After 4 hours of infection, oseltamivir was administered orally for 5 days. The viral loads in the lungs of the group receiving combined treatment were dramatically lower when compared with the viral loads in the lungs of the group receiving oseltamivir alone. HET significantly increased the induction of IL-1β and TNF-α in the lungs of PR8-infected mice and stimulated alveolar macrophage phagocytosis. From these results, we conclude that these functions may be responsible the increased effect on viral load reduction. Here, we show that the combined administration of oseltamivir and HET is very useful for influenza treatment in senescence-accelerated mice.

Heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is a RNA binding protein that plays important role in the biogenesis of mRNA, such as alternative splicing and mRNA stability. We have previously demonstrated that hnRNP A1 has diminished protein levels and shows cytoplasmic accumulation in senescent human diploid fibroblasts. Recent reports showed that p38 MAP kinase (p38 MAPK), a member of the MAP kinase family is necessary and sufficient for the cytoplasmic accumulation of hnRNP A1 by stress stimuli such as osmotic shock. p38 MAP kinase has been shown to be involved in cell proliferation and the induction of senescence in response to extracellular stimuli. However, the relationship between hnRNP A1 and p38 MAPK and the roles of hnRNP A1 in cellularsenescence have not yet been elucidated. Here we show that hnRNP A1 forms a complex with phospho-p38 MAPK in vivo. Inhibition of p38 MAPK activity with SB203580 elevated hnRNP A1 protein levels and prohibited the cytoplasmic accumulation of the protein, but not hnRNP A2, in senescent cells. The phosphorylation level of hnRNP A1 was elevated in senescent cells. Reduction of hnRNP A1 and A2 levels by siRNA transfection induced a senescence-like morphology and elevated the level of F-actin, a marker of senescence. These results suggest that the expression levels and subcellular distribution of hnRNP A1 are regulated in a p38 MAPK-dependent manner, probably via its phosphorylation. Our results also suggest that hnRNP A2 in addition to hnRNP A1 may play a role in establishing the senescence phenotype. PMID:19430204

Although there were considerable advances in the anti-aging medical field, it is short of therapeutic drug for anti-aging. Mounting evidence indicates that the immunosenescence is the key physiopathological mechanism of aging. This study showed the treatment of LW-AFC, an herbal medicine, decreased the grading score of senescence, increased weight, prolonged average life span and ameliorated spatial memory impairment in 12- and 24-month-old senescenceaccelerated mouse resistant 1 (SAMR1) strain. And these anti-aging effects of LW-AFC were more excellent than melatonin. The administration of LW-AFC enhanced ConA- and LPS-induced splenocyte proliferation in aged SAMR1 mice. The treatment of LW-AFC not only reversed the decreased the proportions of helper T cells, suppressor T cells and B cells, the increased regulatory T cells in the peripheral blood of old SAMR1 mice, but also could modulate the abnormal secretion of IL-1β, IL-2, IL-6, IL-17, IL-23, GM-CSF, IFN-γ, TNF-α, TNF-β, RANTES, eotaxin, MCP-1, IL-4, IL-5, IL-10 and G-CSF. These data indicated that LW-AFC reversed the immunosenescence status by restoring immunodeficiency and decreasing chronic inflammation and suggested LW-AFC may be an effective anti-aging agent. PMID:27105505

Although there were considerable advances in the anti-aging medical field, it is short of therapeutic drug for anti-aging. Mounting evidence indicates that the immunosenescence is the key physiopathological mechanism of aging. This study showed the treatment of LW-AFC, an herbal medicine, decreased the grading score of senescence, increased weight, prolonged average life span and ameliorated spatial memory impairment in 12- and 24-month-old senescenceaccelerated mouse resistant 1 (SAMR1) strain. And these anti-aging effects of LW-AFC were more excellent than melatonin. The administration of LW-AFC enhanced ConA- and LPS-induced splenocyte proliferation in aged SAMR1 mice. The treatment of LW-AFC not only reversed the decreased the proportions of helper T cells, suppressor T cells and B cells, the increased regulatory T cells in the peripheral blood of old SAMR1 mice, but also could modulate the abnormal secretion of IL-1β, IL-2, IL-6, IL-17, IL-23, GM-CSF, IFN-γ, TNF-α, TNF-β, RANTES, eotaxin, MCP-1, IL-4, IL-5, IL-10 and G-CSF. These data indicated that LW-AFC reversed the immunosenescence status by restoring immunodeficiency and decreasing chronic inflammation and suggested LW-AFC may be an effective anti-aging agent.

Senescence is not only an important developmental process, but also a responsive regulation to abiotic and biotic stress for plants. Stay-green protein plays crucial roles in plant senescence and chlorophyll degradation. However, the underlying mechanisms were not well-studied, particularly in non-model plants. In this study, a novel stay-green gene, ZjSGR, was isolated from Zoysia japonica. Subcellular localization result demonstrated that ZjSGR was localized in the chloroplasts. Quantitative real-time PCR results together with promoter activity determination using transgenic Arabidopsis confirmed that ZjSGR could be induced by darkness, ABA and MeJA. Its expression levels could also be up-regulated by natural senescence, but suppressed by SA treatments. Overexpression of ZjSGR in Arabidopsis resulted in a rapid yellowing phenotype; complementary experiments proved that ZjSGR was a functional homolog of AtNYE1 from Arabidopsis thaliana. Over expression of ZjSGR accelerated chlorophyll degradation and impaired photosynthesis in Arabidopsis. Transmission electron microscopy observation revealed that overexpression of ZjSGR decomposed the chloroplasts structure. RNA sequencing analysis showed that ZjSGR could play multiple roles in senescence and chlorophyll degradation by regulating hormone signal transduction and the expression of a large number of senescence and environmental stress related genes. Our study provides a better understanding of the roles of SGRs, and new insight into the senescence and chlorophyll degradation mechanisms in plants. PMID:28018416

Senescence is a tumor suppression mechanism that is induced by several stimuli, including oncogenic signaling and telomere shortening, and controlled by the p53/p21(WAF1) signaling pathway. Recently, a critical role for secreted factors has emerged, suggesting that extracellular signals are necessary for the onset and maintenance of senescence. Conversely, factors secreted by senescent cells may promote tumor growth. By using expression profiling techniques, we searched for secreted factors that were overexpressed in fibroblasts undergoing replicative senescence. We identified WNT16B, a member of the WNT family of secreted proteins. We found that WNT16B is overexpressed in cells undergoing stress-induced premature senescence and oncogene-induced senescence in both MRC5 cell line and the in vivo murine model of K-Ras(V12)-induced senescence. By small interfering RNA experiments, we observed that both p53 and WNT16B are necessary for the onset of replicative senescence. WNT16B expression is required for the full transcriptional activation of p21(WAF1). Moreover, WNT16B regulates activation of the phosphoinositide 3-kinase (PI3K)/AKT pathway. Overall, we identified WNT16B as a new marker of senescence that regulates p53 activity and the PI3K/AKT pathway and is necessary for the onset of replicative senescence.

Oligonol is produced from the oligomerization of polyphenols (typically proanthocyanidin from a variety of fruits such as lychees, grapes, apples, persimmons, etc.) and contains catechin-type monomers and oligomers of proanthocyanidins. The ability of Oligonol to affect infection-dependent eye inflammation, locomotion and longevity in senescence-accelerated prone mice (SAMP8) (a model of senescenceacceleration and geriatric disorders with increased oxidative stress and neuronal deficit) was investigated. Oligonol (60mg/kg) significantly modulated the extent of inflammation scores in the eye of SAMP8 mice. Examination of the mice indicated infection with mouse hepatitis virus and pinworm (Syphacia obvelata) in both males and females and with the intestinal protozoa (trichomonad) in males. A comparison of the two groups (using log-rank test) and the difference in the mean life span between groups (using Student's t-test) indicated significant differences in survival (p=0.043) and the mean life span (p=0.033) in male SAMP8 mice. Oligonol increased the mean life span and this was statistically significant. In the open-field locomotive test, the 7-week-old SAMP8 mice crossed more than 40 partitioned lines in 1min. At 48-week-old control untreated male SAMP8 crossed 2 lines. The Oligonol-treated 48-week-old male SAMP8 mice crossed 17 lines however. The improved locomotive activity was statistically significant even after 36weeks in the Oligonol-treated male SAMP8 but this was not the case throughout the time course of the study in the Oligonol-treated female SAMP8. Thus Oligonol treatment to SAMP8 mice modulated the severity of infection-dependent inflammation, prolonged life-span and significantly improved locomotive activity indicating potential benefit to aging-associated diseases such as Alzheimer's or Parkinson's diseases. This presents potential for further research to define infection-dependent inflammation associated with degenerative conditions and the

Glycosylation is one of the most common eukaryotic post-translational modifications, and aberrant glycosylation has been linked to many diseases. However, glycosylation and glycome analysis is a significantly challenging task. Although several lines of evidence have indicated that protein glycosylation is defective in Alzheimer’s disease (AD), only a few studies have focused on AD glycomics. The etiology of AD is unclear and there are no effective disease-modifying treatments for AD. In this study, we found that the object recognition memory, passive avoidance, and spatial learning and memory of senescence-accelerated mouse prone 8 (SAMP8) strain, an AD animal model, were deficient, and LW-AFC, which was prepared from the traditional Chinese medicine prescription Liuwei Dihuang decoction, showed beneficial effects on the deterioration of cognitive capability in SAMP8 mice. Forty-three and 56 N-glycan were identified in the cerebral cortex and serum of SAMP8 mice, respectively. The N-glycan profile in SAMP8 mice was significantly different from that of senescenceaccelerated mouse resistant 1 (SAMR1) strains, the control of SAMP8 mice. Treatment with LW-AFC modulated the abundance of 21 and 6 N-glycan in the cerebral cortex and serum of SAMP8 mice, respectively. The abundance of (Hex)3(HexNAc)5(Fuc)1(Neu5Ac)1 and (Hex)2(HexNAc)4 decreased in the cerebral cortex and serum of SAMP8 mice compared with SAMR1 mice, decreases that were significantly correlated with learning and memory measures. The administration of LW-AFC could reverse or increase these levels in SAMP8 mice. These results indicated that the effects of LW-AFC on cognitive impairments in SAMP8 mice might be through modulation of N-glycan patterns, and LW-AFC may be a potential anti-AD agent. PMID:28203484

Recently, the ability of polyphenols to reduce the risk of dementia and Alzheimer's disease (AD) has attracted a great deal of interest. In the present study, we investigated the attenuating effects of oligomerised lychee fruit-derived polyphenol (OLFP, also called Oligonol) on early cognitive impairment. Male senescence-accelerated mouse prone 8 (SAMP8) mice (4 months old) were given OLFP (100 mg/kg per d) for 2 months, and then conditioned fear memory testing was conducted. Contextual fear memory, which is considered hippocampus-dependent memory, was significantly impaired in SAMP8 mice compared with non-senescence-accelerated mice. OLFP attenuated cognitive impairment in SAMP8 mice. Moreover, the results of real-time PCR analysis that followed DNA array analysis in the hippocampus revealed that, compared with SAMP8 mice, the mRNA expression of Wolfram syndrome 1 (Wfs1) was significantly higher in SAMP8 mice administered with OLFP. Wfs1 reportedly helps to protect against endoplasmic reticulum (ER) stress, which is thought to be one of the causes for AD. The expression of Wfs1 was significantly up-regulated in NG108-15 neuronal cells by the treatment with OLFP, and the up-regulation was inhibited by the treatment of the cells with a c-Jun N-terminal kinase-specific inhibitor rather than with an extracellular signal-regulated kinase inhibitor. Moreover, OLFP significantly attenuated the tunicamycin-induced expression of the ER stress marker BiP (immunoglobulin heavy chain-binding protein) in the cells. These results suggest that OLFP has an attenuating effect on early cognitive impairment in SAMP8 mice, and diminishes ER stress in neuronal cells.

The importance of store-operated Ca2+ entry (SOCE) and the role of its key molecular regulators, STIM1 and ORAI1, in the development of cancer are emerging. Here, we report an unexpected dual function of SOCE in prostate cancer progression by revealing a decrease in the expression of STIM1 in human hyperplasia and tumor tissues of high histological grade and by demonstrating that STIM1 and ORAI1 inhibit cell growth by arresting the G0/G1 phase and enhancing cell senescence in human prostate cancer cells. In addition, STIM1 and ORAI1 inhibited NF-κB signaling and remodeled the tumor microenvironment by reducing the formation of M2 phenotype macrophages, possibly creating an unfavorable tumor microenvironment and inhibiting cancer development. However, STIM1 also promoted cell migration and the epithelial-to-mesenchymal transition by activating TGF-β, Snail and Wnt/β-Catenin pathways. Thus, our study revealed novel regulatory effects and the mechanisms by which STIM1 affects cell senescence, tumor migration and the tumor microenvironment, revealing that STIM1 has multiple functions in prostate cancer cells. PMID:26257076

ABSTRACT Splice variants of certain genes impact on genetic biodiversity in mammals. The tumor suppressor TP53 gene (encoding p53) plays an important role in the regulation of tumorigenesis in hepatocellular carcinoma (HCC). Δ40p53α is a naturally occurring p53 isoform that lacks the N-terminal transactivation domain, yet little is known about the role of Δ40p53α in the development of HCC. Here, we first report on the role of Δ40p53α in HCC cell lines. In the TP53+/Δ40 cell clones, clonogenic activity and cell survival dramatically decreased, whereas the percentage of senescence-associated β-galactosidase (SA-β-gal)-positive cells and p21 (also known as WAF1, CIP1 and CDKN1A) expression significantly increased. These observations were clearly attenuated in the TP53+/Δ40 cell clones after Δ40p53α knockdown. In addition, exogenous Δ40p53 expression significantly suppressed cell growth in HCC cells with wild-type TP53, and in those that were mutant or null for TP53. Notably, Δ40p53α-induced tumor suppressor activity was markedly attenuated in cells expressing the hot-spot mutant Δ40p53α-R175H, which lacks the transcription factor activity of p53. Moreover, Δ40p53α expression was associated with increased full-length p53 protein expression. These findings enhance the understanding of the molecular pathogenesis of HCC and show that Δ40p53α acts as an important tumor suppressor in HCC cells. PMID:27980070

Chronic obstructive pulmonary disease (COPD) is regarded as a disease of accelerated lung aging. This affliction shows all of the hallmarks of aging, including telomere shortening, cellularsenescence, activation of PI3 kinase-mTOR signaling, impaired autophagy, mitochondrial dysfunction, stem cell exhaustion, epigenetic changes, abnormal microRNA profiles, immunosenescence, and a low-grade chronic inflammation (inflammaging). Many of these pathways are driven by chronic exogenous and endogenous oxidative stress. There is also a reduction in antiaging molecules, such as sirtuins and Klotho, which further accelerate the aging process. COPD is associated with several comorbidities (multimorbidity), such as cardiovascular and metabolic diseases, that share the same pathways of accelerated aging. Understanding these mechanisms has helped identify several novel therapeutic targets, and several drugs and dietary interventions are now in development to treat multimorbidity.

Mutations in CDK4 and its key kinase inhibitor p16(INK4a) have been implicated in the genesis and progression of familial human melanoma. The importance of the CDK4 locus in human cancer first became evident following the identification of a germ line CDK4-Arg24Cys (R24C) mutation, which abolishes the ability of CDK4 to bind to p16(INK4a). To determine the role of the Cdk4(R24C) germ line mutation in the genesis of other cancer types, we introduced the R24C mutation in the Cdk4 locus of mice by using Cre-loxP-mediated "knock-in" technology. Cdk4(R24C/R24C) mouse embryo fibroblasts (MEFs) displayed increased Cdk4 kinase activity resulting in hyperphosphorylation of all three members of the Rb family, pRb, p107, and p130. MEFs derived from Cdk4(R24C/R24C) mice displayed decreased doubling times, escape from replicative senescence, and escape sensitivity to contact-induced growth arrest. These MEFs also exhibited a high degree of susceptibility to oncogene-induced transformation, suggesting that the Cdk4(R24C) mutation can serve as a primary event in the progression towards a fully transformed phenotype. In agreement with the in vitro data, homozygous Cdk4(R24C/R24C) mice developed tumors of various etiology within 8 to 10 months of their life span. The majority of these tumors were found in the pancreas, pituitary, brain, mammary tissue, and skin. In addition, Cdk4(R24C/R24C) mice showed extraordinary susceptibility to carcinogens and developed papillomas within the first 8 to 10 weeks following cutaneous application of the carcinogens 9,10-di-methyl-1,2-benz[a]anthracene (DMBA) and 12-O-tetradecanoylphorbol-13-acetate (TPA). This report formally establishes that the activation of Cdk4 is sufficient to promote cancer in many tissues. The observation that a wide variety of tumors develop in mice harboring the Cdk4(R24C) mutation offers a genetic proof that Cdk4 activation may constitute a central event in the genesis of many types of cancers in addition to melanoma.

All over the world people are surviving into their seventh and later decades of life more frequently today than ever before in human history. Some remain in good health, while others show chronic degenerative conditions (CDCs), frailty, and relatively rapid mortality. Thereafter, multiple factors promoting health and well-being become ever more complex as we age. After attainment of reproductive maturation, many physiological decrements occurring in concert with age reflect both senescent and disease processes, not simply the passage of time. Senescence is a process that begins with DNA, molecules and cells and ultimately terminates in cellular death, loss of organ function, and somatic frailty. These changes are different from benign changes with age that do not alter function. Both differ from the pathological processes represented by disease. Either disease or senescence may be age-related, but neither is age-determined. Disease results from pathological alterations and it affects all age groups. Diseases need not be related to senescence, which includes alterations due to inherent aspects of organismal biology. Distinctions among senescence, aging, and disease blur for the late-life CDCs because, in addition to disease processes, many CDCs are phenotypic manifestations of senescing DNA, organelles, cells, and organs. During earlier epochs of human evolution, greater environmental exposures and fewer cultural buffers likely lead to greater frailty and mortality before senescence progressed greatly, as they still do for most animals. In modern-day settings, culturally patterned behaviors have allowed human frailty to become disconnected somewhat from mortality, unlike non-human species.

Research highlights: {yields} SM22{alpha} overexpression in HepG2 cells leads cells to a growth arrest state, and the treatment of a subclinical dose of {gamma}-radiation or doxorubicin promotes cellularsenescence. {yields} SM22{alpha} overexpression elevates p16{sup INK4a} followed by pRB activation, but there are no effects on p53/p21{sup WAF1/Cip1} pathway. {yields} SM22{alpha}-induced MT-1G activates p16{sup INK4a}/pRB pathway, which promotes cellularsenescence by damaging agents. -- Abstract: Smooth muscle protein 22-alpha (SM22{alpha}) is known as a transformation- and shape change-sensitive actin cross-linking protein found in smooth muscle tissue and fibroblasts; however, its functional role remains uncertain. We reported previously that SM22{alpha} overexpression confers resistance against anti-cancer drugs or radiation via induction of metallothionein (MT) isozymes in HepG2 cells. In this study, we demonstrate that SM22{alpha} overexpression leads cells to a growth arrest state and promotes cellularsenescence caused by treatment with a subclinical dose of {gamma}-radiation (0.05 and 0.1 Gy) or doxorubicin (0.01 and 0.05 {mu}g/ml), compared to control cells. Senescence growth arrest is known to be controlled by p53 phosphorylation/p21{sup WAF1/Cip1} induction or p16{sup INK4a}/retinoblastoma protein (pRB) activation. SM22{alpha} overexpression in HepG2 cells elevated p16{sup INK4a} followed by pRB activation, but did not activate the p53/p21{sup WAF1/Cip1} pathway. Moreover, MT-1G, which is induced by SM22{alpha} overexpression, was involved in the activation of the p16{sup INK4a}/pRB pathway, which led to a growth arrest state and promoted cellularsenescence caused by damaging agents. Our findings provide the first demonstration that SM22{alpha} modulates cellularsenescence caused by damaging agents via regulation of the p16{sup INK4a}/pRB pathway in HepG2 cells and that these effects of SM22{alpha} are partially mediated by MT-1G.

Two intracellular fluorochromes, NAD(P)H and Schiff Bases, provide monitoring of energy metabolism and photoperoxidations. Fluorochrome spectra and topographic distribution are measured in a microspectrofluorometer, pixel by pixel using a CCD. The mitochondrial arrangement of Saccharomyces cerevisie and metabolic activity at nuclear kidney epithelial sites is revealed. A kind of accelerated photoaging results in the accumulation of Schiff pigment. Schiff base emission is red-shifted, and it may be preceded by photo-oxidation of NAD(P)H. UVA production of oxygen radicals and peroxides may influence detoxification, senescence and/or transformation. Besides lysosomes, mitochondrial energy metabolism and ER and Golgi detoxification are open to study as multi-organelle complexes with fluorescent xenobiotics and probes. Melanocytes vs. melanoma cells in culture will be investigated using a new compact interferometer for Fourier coding of both emission and excitation spectra. Surprisingly, the photographic method, using the highest sensitivity films, may sometimes produce excellent structural detail. However, for kinetic studies, the CCD, or equivalent, is required. There is good potential for applications in diagnostics and prognostics plus the evaluation of new biopharmeceuticals.

Among a number of non-coding RNAs, role of microRNAs (miRNAs) in cancer cell proliferation, cancer initiation, development and metastasis have been extensively studied and miRNA based therapeutic approaches are being pursued. Prostate cancer (PCa) is a major health concern and several deregulated miRNAs have been described in PCa. miR-212 is differentially modulated in multiple cancers however its function remains elusive. In this study, we found that miR-212 is downregulated in PCa tissues when compared with benign adjacent regions (n = 40). Also, we observed reduced levels of circulatory miR-212 in serum from PCa patients (n = 40) when compared with healthy controls (n = 32). Elucidating the functional role of miR-212, we demonstrate that miR-212 negatively modulates starvation induced autophagy in PCa cells by targeting sirtuin 1 (SIRT1). Overexpression of miR-212 also leads to inhibition of angiogenesis and cellularsenescence. In conclusion, our study indicates a functional role of miR-212 in PCa and suggests the development of miR-212 based therapies. PMID:26439987

Insulin is now considered to be a new drug candidate for treating dementias, such as Alzheimer's disease, whose pathologies are linked to insulin resistance in the brain. Our recent work has clarified that a noncovalent strategy involving cell-penetrating peptides (CPPs) can increase the direct transport of insulin from the nasal cavity into the brain parenchyma. The present study aimed to determine whether the brain insulin level increased by intranasal coadministration of insulin with the CPP penetratin has potential for treating dementia. The pharmacological actions of insulin were investigated at different stages of memory impairment using a senescence-accelerated mouse-prone 8 (SAMP8) model. The results of spatial learning tests suggested that chronic intranasal administration of insulin with l-penetratin to SAMP8 slowed the progression of memory loss in the early stage of memory impairment. However, contrary to expectations, this strategy using penetratin was ineffective in recovering the severe cognitive dysfunction in the progressive stage, which involves brain accumulation of amyloid β (Aβ). Immunohistological examination of hippocampal regions of samples from SAMP8 in the progressive stage suggested that accelerated nose-to-brain insulin delivery had a partial neuroprotective function but unexpectedly increased Aβ plaque deposition in the hippocampus. These findings suggest that the efficient nose-to-brain delivery of insulin combined with noncovalent CPP strategy has different effects on dementia during the mild and progressive stages of cognitive dysfunction.

The senescence-accelerated mouse (SAM) is known to be a murine model for accelerated aging. The SAMP8 strain shows age-related deterioration of learning and memory at an earlier age than control mice (SAMR1). In the present study, we investigated the changes in expressions of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in the brain of SAMP8. In the hippocampus of 10 months old SAMP8, the expression of IL-1 mRNA was significantly elevated in comparison with that of SAMR1. In both strains of SAMs, increases in IL-1beta protein in the brain were observed at 10 months of age compared with 2 and 5 months. The only differences found between the strain in protein levels were at 10 months and were elevations in IL-1beta in the hippocampus and hypothalamus, and in TNF-alpha and IL-6 in the cerebral cortex and the hippocampus in SAMP8 as compared with SAMR1. However, lipopolysaccharide-induced increases in the expression of these cytokines in brain did not differ between SAMP8 and SAMR1. Increases in expression of proinflammatory cytokines in the brain may be involved in the age-related neural dysfunction and/or learning deficiency in SAMP8.

Alzheimer's disease (AD) is a widespread and devastating progressive neurodegenerative disease. Disease-modifying treatments remain beyond reach, and the etiology of the disease is uncertain. Animal model are essential for identifying disease mechanisms and developing effective therapeutic strategies. Research on AD is currently being carried out in rodent models. The most common transgenic mouse model mimics familial AD, which accounts for a small percentage of cases. The senescence-accelerated mouse prone 8 (SAMP8) strain is a spontaneous animal model of accelerated aging. Many studies indicate that SAMP8 mice harbor the behavioral and histopathological signatures of AD, namely AD-like cognitive and behavioral alterations, neuropathological phenotypes (neuron and dendrite spine loss, spongiosis, gliosis and cholinergic deficits in the forebrain), β-amyloid deposits resembling senile plaques, and aberrant hyperphosphorylation of Tau-like neurofibrillary tangles. SAMP8 mice are useful in the development of novel therapies, and many pharmacological agents and approaches are effective in SAMP8 mice. SAMP8 mice are considered a robust model for exploring the etiopathogenesis of sporadic AD and a plausible experimental model for developing preventative and therapeutic treatments for late-onset/age-related AD, which accounts for the vast majority of cases.

Senescence-accelerated mouse prone 8 (SAMP8) and prone 10 (SAMP10) are useful murine model of accelerated aging. SAMP8 shows marked impairment of learning and memory, whereas SAMP10 shows brain atrophy and aging-associated depressive behavior. This study examined the expression of glial cell line-derived neurotrophic factor (GDNF) in SAMP8 and SAMP10 brains, relative to that in SAM resistant 1 (SAMR1) controls, which age normally. Hippocampal GDNF mRNA expression decreased in an age-dependent manner (10- vs 2-month-old animals) in the SAMR1, but not in the SAMP8 or SAMP10 strains. Furthermore, GDNF mRNA expression in 2-month-old SAMP8 and SAMP10 strains was less than in SAMR1 specimens of the same age. The number of surviving neurons in the CA1 region decreased with age in SAMP8 and SAMP10, and also decreased relative to the number of neurons in 10-month-old SAMR1 controls. Immunohistochemistry revealed that cells that were positive for GDNF-like activity in 10-month-old SAMP8 and SAMP10 were diffusely distributed, in part, around the pyramidal cell layer in the hippocampus. These findings suggest that low GDNF expression in young SAMP8 and SAMP10 may be involved in hippocampal dysfunctions, such as age-related learning impairment and neuronal death.

Cellularsenescence can be described as a complex stress response that leads to irreversible cell cycle arrest. This process was originally described as an event that primary cells go through after many passages of cells during cell culture. More recently, cellularsenescence is viewed as a programmed process by which the cell displays a senescence phenotype when exposed to a variety of stresses. Cellularsenescence has been implicated in tumor suppression and aging such that senescence may contribute to both tumor progression and normal tissue repair. Here, we review different forms of cellularsenescence, as well as current biomarkers used to identify senescent cells in vitro and in vivo. Additionally, we highlight the role of senescence-associated long noncoding RNAs (lncRNAs).

Cellularsenescence is often considered a protection mechanism triggered by conditions that impose cellular stress. Continuous proliferation, DNA damaging agents or activated oncogenes are well-known activators of cell senescence. Apart from a characteristic stable cell cycle arrest, this response also involves a proinflammatory phenotype known as senescence-associated secretory phenotype (SASP). This, together with the widely known interference with senescence pathways by some oncoviruses, had led to the hypothesis that senescence may also be part of the host cell response to fight virus. Here, we evaluate this hypothesis using vesicular stomatitis virus (VSV) as a model. Our results show that VSV replication is significantly impaired in both primary and tumor senescent cells in comparison with non-senescent cells, and independently of the stimulus used to trigger senescence. Importantly, we also demonstrate a protective effect of senescence against VSV in vivo. Finally, our results identify the SASP as the major contributor to the antiviral defense exerted by cell senescence in vitro, and points to a role activating and recruiting the immune system to clear out the infection. Thus, our study indicates that cell senescence has also a role as a natural antiviral defense mechanism. PMID:27849057

Estrogen is critical for epiphyseal fusion in both young men and women. In this study, we explored the cellular mechanisms by which estrogen causes this phenomenon. Juvenile ovariectomized female rabbits received either 70 microg/kg estradiol cypionate or vehicle i.m. once a week. Growth plates from the proximal tibia, distal tibia, and distal femur were analyzed after 2, 4, 6, or 8 weeks of treatment. In vehicle-treated animals, there was a gradual senescent decline in tibial growth rate, rate of chondrocyte proliferation, growth plate height, number of proliferative chondrocytes, number of hypertrophic chondrocytes, size of terminal hypertrophic chondrocytes, and column density. Estrogen treatment accelerated the senescent decline in all of these parameters. In senescent growth plates, epiphyseal fusion was observed to be an abrupt event in which all remaining chondrocytes were rapidly replaced by bone elements. Fusion occurred when the rate of chondrocyte proliferation approached zero. Estrogen caused this proliferative exhaustion and fusion to occur earlier. Our data suggest that (i) epiphyseal fusion is triggered when the proliferative potential of growth plate chondrocytes is exhausted; and (ii) estrogen does not induce growth plate ossification directly; instead, estrogen accelerates the programmed senescence of the growth plate, thus causing earlier proliferative exhaustion and consequently earlier fusion.

The widespread environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a potent toxicant that causes significant neurotoxicity. However, the biological events that participate in this process remain largely elusive. In the present study, we demonstrated that TCDD exposure triggered apparent premature senescence in rat pheochromocytoma (PC12) and human neuroblastoma SH-SY5Y cells. Senescence-associated β-galactosidase (SA-β-Gal) assay revealed that TCDD induced senescence in PC12 neuronal cells at doses as low as 10 nM. TCDD led to F-actin reorganization and the appearance of an alternative senescence marker, γ-H2AX foci, both of which are important features of cellularsenescence. In addition, TCDD exposure altered the expression of senescence marker proteins, such as p16, p21 and p-Rb, in both dose- and time-dependent manners. Furthermore, we demonstrated that TCDD promotes mitochondrial dysfunction and the accumulation of cellular reactive oxygen species (ROS) in PC12 cells, leading to the activation of signaling pathways that are involved in ROS metabolism and senescence. TCDD-induced ROS generation promoted significant oxidative DNA damage and lipid peroxidation. Notably, treatment with the ROS scavenger N-acetylcysteine (NAC) markedly attenuated TCDD-induced ROS production, cellular oxidative damage and neuronal senescence. Moreover, we found that TCDD induced a similar ROS-mediated senescence response in human neuroblastoma SH-SY5Y cells. In sum, these results demonstrate for the first time that TCDD induces premature senescence in neuronal cells by promoting intracellular ROS production, supporting the idea that accelerating the onset of neuronal senescence may be an important mechanism underlying TCDD-induced neurotoxic effects.

The senescence-accelerated mouse prone10 (SAMP10) strain, a model of aging, exhibits cognitive impairments and cerebral atrophy. We noticed that SAMP10/TaSlc mice, a SAMP10 substrain, have developed persistent glucosuria over the past few years. In the present study, we characterized SAMP10/TaSlc mice and further identified a spontaneous mutation in the Slc5a2 gene encoding sodium-glucose co-transporter (SGLT) 2. The mean concentration of urine glucose was high in SAMP10/TaSlc mice and increased further with advancing age, whereas other strains of senescence-accelerated mice, including SAMP1/SkuSlc, SAMP6/TaSlc and SAMP8/TaSlc or normal aging control SAMR1/TaSlc mice, exhibited no detectable glucose in urine. SAMP10/TaSlc mice consumed increasing amounts of food and water compared to SAMR1/TaSlc mice, suggesting the compensation of polyuria and the loss of glucose. Oral glucose tolerance tests showed decreased glucose reabsorption in the kidney of SAMP10/TaSlc mice. In addition, blood glucose levels decreased in an age-dependent fashion. The kidney was innately larger than that of control mice with no histological alterations. We examined the expression levels of glucose transporters in the kidney. Among SGLT1, SGLT2, glucose transporter (GLUT) 1 and GLUT2, we found a significant decrease only in the level of SGLT2. DNA sequencing of SGLT2 in SAMP10/TaSlc mice revealed a single nucleotide deletion of guanine at 1236, which resulted in a frameshift mutation that produced a truncated protein. We designate this strain as SAMP10/TaSlc-Slc5a2(slc) (SAMP10-ΔSglt2). Recently, SGLT2 inhibitors have been demonstrated to be effective for the treatment of patients with type 2 diabetes (T2D). SAMP10-ΔSglt2 mice may serve as a unique preclinical model to study the link between aging-related neurodegenerative disorders and T2D.

Background: Amyloid β (Aβ) has been established as a key factor for the pathological changes in the brains of patients with Alzheimer's disease (AD), and cellularsenescence is closely associated with aging and cognitive impairment. However, it remains blurred whether, in the AD brains, Aβ accelerates the neuronal senescence and whether this senescence, in turn, impairs the cognitive function. This study aimed to explore the expression of senescence-associated genes in the hippocampal tissue from young to aged 5XFAD mice and their age-matched wild type (WT) mice to determine whether senescent neurons are present in the transgenic AD mouse model. Methods: The 5XFAD mice and age-matched wild type mice, both raised from 1 to 18 months, were enrolled in the study. The senescence-associated genes in the hippocampus were analyzed and differentially expressed genes (DEGs) were screened by quantitative real-time polymerase chain reaction. Cognitive performance of the mice was evaluated by Y-maze and Morris water maze tests. Oligomeric Aβ (oAβ) (1–42) was applied to culture primary neurons to simulate the in vivo manifestation. Aging-related proteins were detected by Western blotting analysis and immunofluorescence. Results: In 5XFAD mice, of all the DEGs, the senescence-associated marker p16 was most significantly increased, even at the early age. It was mainly localized in neurons, with a marginal expression in astrocytes (labeled as glutamine synthetase), nil expression in activated microglia (labeled as Iba1), and negatively correlated with the spatial cognitive impairments of 5XFAD mice. oAβ (1–42) induced the production of senescence-related protein p16, but not p53 in vitro, which was in line with the in vivo manifestation. Conclusions: oAβ-accelerated neuronal senescence may be associated with the cognitive impairment in 5XFAD mice. Senescence-associated marker p16 can serve as an indicator to estimate the cognitive prognosis for AD population. PMID

This study examines the cellular automata traffic flow model, which considers the heterogeneity of vehicle acceleration and the delay probability of vehicles. Computer simulations are used to identify three typical phases in the model: free-flow, synchronized flow, and wide moving traffic jam. In the synchronized flow region of the fundamental diagram, the low and high velocity vehicles compete with each other and play an important role in the evolution of the system. The analysis shows that there are two types of bistable phases. However, in the original Nagel and Schreckenberg cellular automata traffic model, there are only two kinds of traffic conditions, namely, free-flow and traffic jams. The synchronized flow phase and bistable phase have not been found.

Alzheimer's disease (AD), the most common age-dependent neurodegenerative disorder, produces a progressive decline in cognitive function. The metabolic mechanism of AD has emerged in recent years. In this study, we used multivariate analyses of gas chromatography-mass spectrometry measurements to determine that learning and retention-related metabolic profiles are altered during aging in the hippocampus of the senescence-accelerated mouse prone 8 (SAMP8). Alterations in 17 metabolites were detected in mature and aged mice compared to young mice (13 decreased and 4 increased metabolites), including metabolites related to dysfunctional lipid metabolism (significantly increased cholesterol, oleic acid, and phosphoglyceride levels), decreased amino acid (alanine, serine, glycine, aspartic acid, glutamate, and gamma-aminobutyric acid), and energy-related metabolite levels (malic acid, butanedioic acid, fumaric acid, and citric acid), and other altered metabolites (increased N-acetyl-aspartic acid and decreased pyroglutamic acid, urea, and lactic acid) in the hippocampus. All of these alterations indicated that the metabolic mechanisms of age-related cognitive impairment in SAMP8 mice were related to multiple pathways and networks. Lipid metabolism, especially cholesterol metabolism, appears to play a distinct role in the hippocampus in AD. PMID:24284365

Tiantai No. 1, a Chinese medicine predominantly composed of powdered Rhizoma Gastrodiae, Radix Ginseng, and Ginkgo leaf at a ratio of 2:1:2 and dissolved in pure water, is neuroprotective in animal models of various cognitive disorders, but its molecular mechanism remains unclear. We administered Tiantai No. 1 intragastrically to senescence-accelerated mouse prone 8 (SAMP8) mice (a model of Alzheimer's disease) at doses of 50, 100 or 150 mg/kg per day for 8 weeks and evaluated their behavior in the Morris water maze and expression of Alzheimer's disease-related proteins in the brain. Tiantai No. 1 shortened the escape latency in the water maze training trials, and increased swimming time in the target quadrant during the spatial probe test, indicating that Tiantai No. 1 improved learning and memory in SAMP8 mice. Immunohistochemistry revealed that Tiantai No. 1 restored the proliferation potential of Ki67-positive cells in the hippocampus. In addition, mice that had received Tiantai No. 1 had fewer astrocytes, and less accumulation of amyloid-beta and phosphorylated tau. These results suggest that Tiantai No. 1 is neuroprotective in the SAMP8 mouse model of Alzheimer's disease and acts by restoring neuronal number and proliferation potential in the hippocampus, decreasing astrocyte infiltration, and reducing the accumulation of amyloid-beta and phosphorylated tau.

We previously showed that mitochondria-targeted antioxidant SkQ1 (plastoquinonyl-decyltriphenylphosphonium) at nanomolar concentrations is capable of preventing and slowing down some cerebral dysfunctions in accelerated-senescence OXYS rats. Here we demonstrate that OXYS rats develop behavior, learning, and memory deficits against a background of neurodegeneration signs detected by magnetic resonance tomography and amyloid-β (Aβ) pathology similar to those seen in Alzheimer's disease (AD). Long-term treatment with SkQ1 (250 nmol/kg body weight daily from the age of 1.5 to 23 months) reduced the age-related alterations in behavior and spatial memory deficit in Morris water maze in OXYS and Wistar rats. Furthermore, this is the first report that SkQ1 treatment slows down pathological accumulation of AβPP, Aβ, and hyperphosphorylation of tau-protein in OXYS rats, as well as age-dependent changes in healthy Wistar rats. Our results support the possibility of using the OXYS strain as a rat model of AD-like pathology. It seems probable that the mitochondria-targeted antioxidant SkQ1 can be a good prophylactic strategy to maintain brain health and to treat AD.

The decline in physical performance with increasing age is a crucial problem in our aging society. We examined the effects of resveratrol, a natural polyphenolic compound present in grapes, in combination with habitual exercise on the aging-associated decline in physical performance in senescence-accelerated prone mice (SAMP1). The endurance capacity of SAMP1 mice undergoing an exercise regimen (SAMP1-Ex) decreased over 12 weeks whereas that of SAMP1 mice fed 0.2% (w/w) resveratrol along with exercise (SAMP1-ExRes) remained significantly higher. In the SAMP1-ExRes group, there was a significant increase in oxygen consumption and skeletal muscle mRNA levels of mitochondrial function-related enzymes. These results suggest that the intake of resveratrol, together with habitual exercise, is beneficial for suppressing the aging-related decline in physical performance and that these effects are attributable, at least in part, to improved mitochondrial function in skeletal muscle.

Summary Cellularsenescence is an irreversible arrest of cell proliferation at the G1 stage of the cell cycle in which cells become refractory to growth stimuli. Senescence is a critical and potent defense mechanism that mammalian cells have to suppress tumors. While there are many ways to induce a senescence response, oncogene-induced senescence (OIS) remains key to inhibiting progression of cells that have acquired oncogenic mutations. In primary cells in culture, OIS induces a set of measurable phenotypic and behavioral changes, in addition to cell cycle exit. Senescence-associated β-Galactosidase (SA-β-Gal) activity is a main hallmark of senescent cells, along with morphological changes that may depend on the oncogene that is activated, or on the primary cell type. Characteristic cellular changes of senescence include increased size, flattening, multi-nucleation, and extensive vacuolation. At the molecular level, tumor suppressor genes such as p53 and p16INK4a may play a role in initiation or maintenance of OIS. Activation of a DNA damage response and a senescence-associated secretory phenotype could delineate the onset of senescence. Despite advances in our understanding of how OIS suppresses some tumor types, the in vivo role of OIS in melanocytic nevi and melanoma remains poorly understood and not validated. In an effort to stimulate research in this field, we review in this chapter the known markers of senescence and provide experimental protocols for their identification by immunofluorescent staining in melanocytic nevi and malignant melanoma. PMID:27812879

Atherosclerosis is a disease of ageing in that its incidence and prevalence increase with age. However, atherosclerosis is also associated with biological ageing, manifest by a number of typical hallmarks of ageing in the atherosclerotic plaque. Thus, accelerated biological ageing may be superimposed on the effects of chronological ageing in atherosclerosis. Tissue ageing is seen in all cells that comprise the plaque, but particularly in vascular smooth muscle cells (VSMCs). Hallmarks of ageing include evidence of cell senescence, DNA damage (including telomere attrition), mitochondrial dysfunction, a pro-inflammatory secretory phenotype, defects in proteostasis, epigenetic changes, deregulated nutrient sensing, and exhaustion of progenitor cells. In this model, initial damage to DNA (genomic, telomeric, mitochondrial and epigenetic changes) results in a number of cellular responses (cellularsenescence, deregulated nutrient sensing and defects in proteostasis). Ultimately, ongoing damage and attempts at repair by continued proliferation overwhelm reparative capacity, causing loss of specialised cell functions, cell death and inflammation. This review summarises the evidence for accelerated biological ageing in atherosclerosis, the functional consequences of cell ageing on cells comprising the plaque, and the causal role that VSMC senescence plays in atherogenesis.

Bone marrow-derived cells enter the brain in a non-inflammatory condition through the attachments of choroid plexus and differentiate into ramified myeloid cells. Neurodegenerative conditions may be associated with altered immune-brain interaction. The senescence-accelerated mouse prone 10 (SAMP10) undergoes earlier onset neurodegeneration than C57BL/6 (B6) strain. We hypothesized that the dynamics of immune cells migrating from the bone marrow to the brain is perturbed in SAMP10 mice. We created 4 groups of radiation chimeras by intra-bone marrow-bone marrow transplantation using 2-month-old (2 mo) and 10 mo SAMP10 and B6 mice as recipients with GFP transgenic B6 mice as donors, and analyzed histologically 4 months later. In the [B6 → 10 mo SAMP10] chimeras, more ramified marrow-derived cells populated a larger number of discrete brain regions than the other chimeras, especially in the diencephalon. Multiplex cytokine assays of the diencephalon prepared from non-treated 3 mo and 12 mo SAMP10 and B6 mice revealed that 12 mo SAMP10 mice exhibited higher tissue concentrations of CXCL1, CCL11, G-CSF, CXCL10 and IL-6 than the other groups. Immunohistologically, choroid plexus epithelium and ependyma produced CXCL1, while astrocytic processes in the attachments of choroid plexus expressed CCL11 and G-CSF. The median eminence produced CXCL10, hypothalamic neurons G-CSF and tanycytes CCL11 and G-CSF. These brain cytokine profile changes in 12 mo SAMP10 mice were likely to contribute to acceleration of the dynamics of marrow-derived cells to the diencephalon. Further studies on the functions of ramified marrow-derived myeloid cells would enhance our understanding of the brain-bone marrow interaction.

Aging is considered to be a functional retardation of continuous xenobiotic responses over a lifetime after the developmental period; thus, the effects of ionizing radiation over a lifetime may be somewhat accounted for by a modifier of aging effects. This study was conducted to evaluate the possible/synergic effects of radiation during aging by determining cell-cycle parameters of hematopoietic stem cells/hematopoietic progenitor cells (HSCs/HPCs), such as the percent of cells in cycling, the generation doubling time, and the cumulative cycling-cell fraction, by bromodeoxyuridine-ultraviolet assay, which enables the determination of their cycling capacity in vivo. Colony-forming progenitor cells, such as colony-forming unit (CFU)-granulocyte/macrophage (GM), CFU in the spleen on day 9 (CFU-S9), and CFU-S on day 13 (CFU-S13) for mature, less mature, and immature HPCs, respectively, were evaluated in young and old mice (6 weeks and 21 months of age, respectively) with or without 2-Gy whole-body irradiation and a 4-week recovery period. Then, cell-cycle parameters were evaluated and compared. As a result, the generation doubling time of all types of HPC was prolonged by the irradiation in both young and old mouse groups, except that of CFU-S13 in old mice, which showed acceleration of the cell cycle following the irradiation. In addition, only CFU-S13 in irradiated old mice showed a significant increase in the cumulative cycling-cell-fraction ratio. Significant changes due to the effects of aging and irradiation on HPCs were observed only in the immature HPCs, i.e., the cell cycle of immature HPCs was suppressed by aging without irradiation and was, in contrast, accelerated as the cells recovered from radiation-induced damage. This suggests that the mechanisms of peripheral blood recovery after 2-Gy whole-body irradiation are markedly different between young and old mice, although 21-month-old mice showed almost the same level of recovery as the young mice.

Recent studies in human fibroblasts have provided a new general paradigm of tumor suppression according to which oncogenic signaling produces DNA damage and this, in turn, results in ATM/p53-dependent cellularsenescence. Here, we have tested this model in a variety of murine experimental systems. Overexpression of oncogenic Ras in murine fibroblasts efficiently induced senescence but this occurred in the absence of detectable DNA damage signaling, thus suggesting a fundamental difference between human and murine cells. Moreover, lung adenomas initiated by endogenous levels of oncogenic K-Ras presented abundant senescent cells, but undetectable DNA damage signaling. Accordingly, K-Ras-driven adenomas were also senescent in Atm-null mice, and the tumorigenic progression of these lesions was only modestly accelerated by Atm-deficiency. Finally, we have examined chemically-induced fibrosarcomas, which possess a persistently activated DNA damage response and are highly sensitive to the activity of p53. We found that the absence of Atm favored genomic instability in the resulting tumors, but did not affect the persistent DNA damage response and did not impair p53-dependent tumor suppression. All together, we conclude that oncogene-induced senescence in mice may occur in the absence of a detectable DNA damage response. Regarding murine Atm, our data suggest that it plays a minor role in oncogene-induced senescence or in p53-dependent tumor suppression, being its tumor suppressive activity probably limited to the maintenance of genomic stability. PMID:19421407

Most normal human cells have a finite replicative capacity and eventually undergo cellularsenescence, whereby cells cease to proliferate. Cellularsenescence is also induced by various stress signals, such as those generated by oncogenes, DNA damage, hyperproliferation, and an oxidative environment. Cellularsenescence is well established as an intrinsic tumor suppressive mechanism. Recent progress concerning senescence research has revealed that cellularsenescence occurs in vivo and that, unexpectedly, it has a very complex role in tissue repair, promoting tumor progression and aging via the secretion of various cytokines, growth factors, and enzymes. Therefore, the importance of biomarkers for cellularsenescence has greatly increased. In 1995, we described the "senescence-associated β galactosidase" (SA-βgal) biomarker, which conveniently identifies individual senescent cells in vitro and in vivo. Here, we describe an updated protocol for the detection of cell senescence based on this widely used biomarker, which contributed to recent advances in senescence, aging and cancer research. We provide an example of detecting SA-βgal together with other senescence markers and a proliferation marker, EdU, in single cells.

Amyloid β-peptide (Aβ) plays a central role in the pathophysiology of Alzheimer's disease (AD) through the induction of oxidative stress. This peptide is produced by proteolytic cleavage of amyloid precursor protein (APP) by the action of β- and γ-secretases. Previous studies demonstrated that reduction of Aβ, using an antisense oligonucleotide (AO) directed against the Aβ region of APP, reduced oxidative stress-mediated damage and prevented or reverted cognitive deficits in senescence-accelerated prone mice (SAMP8), a useful animal model for investigating the events related to Aβ pathology and possibly to the early phase of AD. In the current study, aged SAMP8 were treated by AO directed against PS-1, a component of the γ-secretase complex, and tested for learning and memory in T-maze foot shock avoidance and novel object recognition. Brain tissue was collected to identify the decrease of oxidative stress and to evaluate the proteins that are differently expressed and oxidized after the reduction in free radical levels induced by Aβ. We used both expression proteomics and redox proteomics approaches. In brain of AO-treated mice a decrease of oxidative stress markers was found, and the proteins identified by proteomics as expressed differently or nitrated are involved in processes known to be impaired in AD. Our results suggest that the treatment with AO directed against PS-1 in old SAMP8 mice reverses learning and memory deficits and reduces Aβ-mediated oxidative stress with restoration to the normal condition and identifies possible pharmacological targets to combat this devastating dementing disease.

Vitamin K2 (VK2) has been used as a therapeutic agent for osteoporosis, since it has been suggested to be able to reduce the frequency of fractures by improving bone quality; however, bone turnover is strictly regulated by various cytokines and hormones. In the present study, the effect of menaquinone-4 (MK-4) on bone turnover was investigated using the senescence-accelerated mouse prone 6 (SAMP6) strain. Since water-immersion restraint stress (WRS) causes a significant decrease in bone mineral density (BMD), WRS was used as the bone resorption model in the SAMP6 strain. Six-week-old SAMP6 male mice were divided into the following three groups: Control, WRS and WRS + MK-4. WRS was performed for 6 h per day, 5 times a week, for 4 weeks. Following WRS, MK-4 (30 mg/kg) was injected subcutaneously 3 times a week for 4 weeks. No growth retardation was observed in the WRS groups as compared with the control group. In the WRS groups, the BMD was significantly lower than that in the control group. The levels of bone formation and resorption markers were increased in the WRS groups, indicating that WRS reduced the BMD by promoting high bone turnover. A bone histomorphometrical examination showed that the trabecular (Tb) bone mass in the secondary spongiosa at the distal femur was significantly reduced in the WRS mice, and this reduction was abrogated by MK-4 treatment. Specifically, the Tb bone reduction was caused by the activation of osteoclasts (Ocs), and Oc activity was suppressed by MK-4. The number of osteoblasts and the mineral apposition rate were significantly increased in the WRS and WRS + MK-4 mice, suggesting that WRS triggered a significantly higher mineral apposition rate. These results indicate that MK-4 can induce recovery from the bone mineral loss caused by WRS treatment. Further studies are required to clarify the association between bone quality and MK-4. PMID:26622403

Sarcopenia-related falls and fall-related injuries in community-dwelling elderly people garnered more and more interest in recent years. Low-magnitude high-frequency vibration (LMHFV) was proven beneficial to musculoskeletal system and recommended for sarcopenia treatment. This study aimed to evaluate the effects of LMHFV on the sarcopenic animals and explore the mechanism of the stimulatory effects. Senescence-accelerated mouse P8 (SAMP8) mice at month 6 were randomized into control (Ctrl) and vibration (Vib) groups and the mice in the Vib group were given LMHFV (0.3 g, 20 min/day, 5 days/week) treatment. At months 0, 1, 2, 3, and 4 post-treatment, muscle mass, structure, and function were assessed. The potential proliferation capacity of the muscle was also evaluated by investigating satellite cells (SCs) pool and serum myostatin expression. At late stage, the mice in the Vib group showed higher muscle strength (month 4, p = 0.028). Generally, contractibility was significantly improved by LMHFV (contraction time [CT], p = 0.000; half-relaxation time [RT50], p = 0.000). Enlarged cross-sectional area of fiber type IIA was observed in the Vib group when compared with Ctrl group (p = 0.000). No significant difference of muscle mass was observed. The promotive effect of LMHFV on myoregeneration was reflected by suppressed SC pool reduction (month 3, p = 0.000; month 4, p = 0.000) and low myostatin expression (p = 0.052). LMHFV significantly improved the structural and functional outcomes of the skeletal muscle, hence retarding the progress of sarcopenia in SAMP8. It would be a good recommendation for prevention of the diseases related to skeletal muscle atrophy.

Cellularsenescence suppresses cancer by arresting cell proliferation, essentially permanently, in response to oncogenic stimuli, including genotoxic stress. We modified the use of antibody arrays to provide a quantitative assessment of factors secreted by senescent cells. We show that human cells induced to senesce by genotoxic stress secrete myriad factors associated with inflammation and malignancy. This senescence-associated secretory phenotype (SASP) developed slowly over several days and only after DNA damage of sufficient magnitude to induce senescence. Remarkably similar SASPs developed in normal fibroblasts, normal epithelial cells, and epithelial tumor cells after genotoxic stress in culture, and in epithelial tumor cells in vivo after treatment of prostate cancer patients with DNA-damaging chemotherapy. In cultured premalignant epithelial cells, SASPs induced an epithelial-mesenchyme transition and invasiveness, hallmarks of malignancy, by a paracrine mechanism that depended largely on the SASP factors interleukin (IL)-6 and IL-8. Strikingly, two manipulations markedly amplified, and accelerated development of, the SASPs: oncogenic RAS expression, which causes genotoxic stress and senescence in normal cells, and functional loss of the p53 tumor suppressor protein. Both loss of p53 and gain of oncogenic RAS also exacerbated the promalignant paracrine activities of the SASPs. Our findings define a central feature of genotoxic stress-induced senescence. Moreover, they suggest a cell-nonautonomous mechanism by which p53 can restrain, and oncogenic RAS can promote, the development of age-related cancer by altering the tissue microenvironment.

AMPK activation is beneficial for cellular homeostasis and senescence prevention. However, the molecular events involved in AMPK activation are not well defined. In this study, we addressed the mechanism underlying the protective effect of AMPK on oxidative stress-induced senescence. The results showed that AMPK was inactivated in senescent cells. However, pharmacological activation of AMPK by metformin and berberine significantly prevented the development of senescence and, accordingly, inhibition of AMPK by Compound C was accelerated. Importantly, AMPK activation prevented hydrogen peroxide-induced impairment of the autophagic flux in senescent cells, evidenced by the decreased p62 degradation, GFP-RFP-LC3 cancellation, and activity of lysosomal hydrolases. We also found that AMPK activation restored the NAD(+) levels in the senescent cells via a mechanism involving mostly the salvage pathway for NAD(+) synthesis. In addition, the mechanistic relationship of autophagic flux and NAD(+) synthesis and the involvement of mTOR and Sirt1 activities were assessed. In summary, our results suggest that AMPK prevents oxidative stress-induced senescence by improving autophagic flux and NAD(+) homeostasis. This study provides a new insight for exploring the mechanisms of aging, autophagy and NAD(+) homeostasis, and it is also valuable in the development of innovative strategies to combat aging.

Cellularsenescence suppresses cancer by arresting cell proliferation, essentially permanently, in response to oncogenic stimuli, including genotoxic stress. We modified the use of antibody arrays to provide a quantitative assessment of factors secreted by senescent cells. We show that human cells induced to senesce by genotoxic stress secrete myriad factors associated with inflammation and malignancy. This senescence-associated secretory phenotype (SASP) developed slowly over several days and only after DNA damage of sufficient magnitude to induce senescence. Remarkably similar SASPs developed in normal fibroblasts, normal epithelial cells, and epithelial tumor cells after genotoxic stress in culture, and in epithelial tumor cells in vivo after treatment of prostate cancer patients with DNA-damaging chemotherapy. In cultured premalignant epithelial cells, SASPs induced an epithelial–mesenchyme transition and invasiveness, hallmarks of malignancy, by a paracrine mechanism that depended largely on the SASP factors interleukin (IL)-6 and IL-8. Strikingly, two manipulations markedly amplified, and accelerated development of, the SASPs: oncogenic RAS expression, which causes genotoxic stress and senescence in normal cells, and functional loss of the p53 tumor suppressor protein. Both loss of p53 and gain of oncogenic RAS also exacerbated the promalignant paracrine activities of the SASPs. Our findings define a central feature of genotoxic stress-induced senescence. Moreover, they suggest a cell-nonautonomous mechanism by which p53 can restrain, and oncogenic RAS can promote, the development of age-related cancer by altering the tissue microenvironment. PMID:19053174

Accelerator mass spectrometry (AMS) is a highly sensitive analytical methodology used to quantify the content of radioisotopes, such as {sup 14}C, in a sample. The primary goals of this work were to demonstrate the utility of AMS in determining cellular [{sup 14}C]doxorubicin (DOX) concentrations and to develop a sensitive assay that is superior to high performance liquid chromatography (HPLC) for the quantification of DOX at the tumor level. In order to validate the superior sensitivity of AMS versus HPLC with fluorescence detection, we performed three studies comparing the cellular accumulation of DOX: one in vitro cell line study, and two in vivo xenograft mouse studies. Using AMS, we quantified cellular DOX content up to 4 hours following in vitro exposure at concentrations ranging from 0.2 pg/ml (345 fM) to 2 {micro}g/ml (3.45 {micro}M) [{sup 14}C]DOX. The results of this study show that, compared to standard fluorescence-based HPLC, the AMS method was over five orders of magnitude more sensitive. Two in vivo studies compared the sensitivity of AMS to HPLC using a nude mouse xenograft model in which breast cancer cells were implanted subcutaneously. After sufficiently large tumors formed, DOX was administered intravenously at two dose levels. Additionally, we tested the AMS method in a nude mouse xenograft model of multidrug resistance (MDR) in which each mouse was implanted with both wild type and MDR+ cells on opposite flanks. The results of the second and third studies showed that DOX concentrations were significantly higher in the wild type tumors compared to the MDR+ tumors, consistent with the MDR model. The extreme sensitivity of AMS should facilitate similar studies in humans to establish target site drug delivery and to potentially determine the optimal treatment dose and regimen.

We describe the basic tenets of the current concepts of cancer biology, and review the recent advances on the suppressor role of senescence in tumor growth and the breakdown of this barrier during the origin of tumor growth. Senescence phenotype can be induced by (1) telomere attrition-induced senescence at the end of the cellular mitotic life span (MLS*) and (2) also by replication history-independent, acceleratedsenescence due to inadvertent activation of oncogenes or by exposure of cells to genotoxins. Tumor suppressor genes p53/pRB/p16INK4A and related senescence checkpoints are involved in effecting the onset of senescence. However, senescence as a tumor suppressor mechanism is a leaky process and senescent cells with mutations or epimutations in these genes escape mitotic catastrophe-induced cell death by becoming polyploid cells. These polyploid giant cells, before they die, give rise to several cells with viable genomes via nuclear budding and asymmetric cytokinesis. This mode of cell division has been termed neosis and the immediate neotic offspring the Raju cells. The latter inherit genomic instability and transiently display stem cell properties in that they differentiate into tumor cells and display extended, but, limited MLS, at the end of which they enter senescent phase and can undergo secondary/tertiary neosis to produce the next generation of Raju cells. Neosis is repeated several times during tumor growth in a non-synchronized fashion, is the mode of origin of resistant tumor growth and contributes to tumor cell heterogeneity and continuity. The main event during neosis appears to be the production of mitotically viable daughter genome after epigenetic modulation from the non-viable polyploid genome of neosis mother cell (NMC). This leads to the growth of resistant tumor cells. Since during neosis, spindle checkpoint is not activated, this may give rise to aneuploidy. Thus, tumor cells also are destined to die due to senescence, but may escape

Long-term consumption of artificial sweeteners (AS) has been the recent focus of safety concerns. However, the potential risk of the AS in cardiovascular disease and lipoprotein metabolism has not been investigated sufficiently. We compared the influence of AS (aspartame, acesulfame K, and saccharin) and fructose in terms of functional and structural correlations of apolipoprotein (apo) A-I and high-density lipoproteins (HDL), which have atheroprotective effects. Long-term treatment of apoA-I with the sweetener at physiological concentration (3 mM for 168 h) resulted in loss of antioxidant and phospholipid binding activities with modification of secondary structure. The AS treated apoA-I exhibited proteolytic cleavage to produce 26 kDa-fragment. They showed pro-atherogenic properties in acetylated LDL phagocytosis of macrophages. Each sweetener alone or sweetener-treated apoA-I caused acceleratedsenescence in human dermal fibroblasts. These results suggest that long-term consumption of AS might accelerate atherosclerosis and senescence via impairment of function and structure of apoA-I and HDL.

E-cadherin (CDH1) is a cell adhesion molecule that coordinates key morphogenetic processes regulating cell growth, cell proliferation, and apoptosis. Loss of CDH1 is a trademark of the cellular event epithelial to mesenchymal transition, which increases the metastatic potential of malignant cells. PTEN is a tumor-suppressor gene commonly mutated in many human cancers, including endometrial cancer. In the mouse uterus, ablation of Pten induces epithelial hyperplasia, leading to endometrial carcinomas. However, loss of Pten alone does not affect longevity until around 5 mo. Similarly, conditional ablation of Cdh1 alone does not predispose mice to cancer. In this study, we characterized the impact of dual Cdh1 and Pten ablation (Cdh1(d/d) Pten(d/d)) in the mouse uterus. We observed that Cdh1(d/d) Pten(d/d) mice died at Postnatal Days 15-19 with massive blood loss. Their uteri were abnormally structured with curly horns, disorganized epithelial structure, and increased cell proliferation. Co-immunostaining of KRT8 and ACTA2 showed invasion of epithelial cells into the myometrium. Further, the uteri of Cdh1(d/d) Pten(d/d) mice had prevalent vascularization in both the endometrium and myometrium. We also observed reduced expression of estrogen and progesterone receptors, loss of cell adherens, and tight junction molecules (CTNNB1 and claudin), as well as activation of AKT in the uteri of Cdh1(d/d) Pten(d/d) mice. However, complex hyperplasia was not found in the uteri of Cdh1(d/d) Pten(d/d) mice. Collectively, these findings suggest that ablation of Pten with Cdh1 in the uterus acceleratescellular invasiveness and angiogenesis and causes early death.

Eldecalcitol (ELD), a 2β-hydroxypropyloxy derivative of 1α,25(OH)2D3, is a potent inhibitor of bone resorption that has demonstrated a greater effect at reducing the risk of fracture in osteoporotic patients than alfacalcidol (ALF). In the present study, we used the senescence-accelerated mouse strain P6 (SAM/P6), which has low bone mass caused by osteoblast dysfunction, to evaluate the effect of ELD on cortical bone in comparison with ALF. Four-month-old SAM/P6 mice were given either ELD (0.025 or 0.05μg/kg) or ALF (0.2 or 0.4μg/kg) by oral gavage 5 times/week for 6 weeks. Both ELD and ALF increased serum calcium (Ca) in a dose-dependent manner. Serum Ca levels in the ELD 0.05μg/kg group were comparable to those of the ALF 0.2μg/kg group. ELD 0.05μg/kg significantly improved the bone biomechanical properties of the femur compared with the vehicle control group (p<0.001) and the ALF 0.2μg/kg group (p<0.05) evaluated by 3-point bending test. The cortical area of the mid-femur in the ELD 0.05μg/kg group but not the ALF 0.2μg/kg group was significantly higher than those of the vehicle control group (p<0.001). Bone histomorphometry revealed that in the femoral endocortical surface, the suppression of bone resorption parameters (N.Oc/BS) and bone formation parameters (MS/BS) by ELD (0.05μg/kg) was greater than that by ALF (0.2μg/kg). In contrast, in the femoral periosteal surface, ELD 0.05μg/kg significantly increased bone formation parameters (BFR/BS, MS/BS) compared with the vehicle control group (p<0.05, p<0.01, respectively), whereas ALF 0.2μg/kg did not alter these parameters. These results indicate that ELD improved the biomechanical properties of femoral cortical bone not only by inhibiting endocortical bone resorption but also by stimulating the periosteal bone formation in SAM/P6 mice. This article is part of a Special Issue entitled '16th Vitamin D Workshop'.

The identification of the physical mechanism(s) by which cells can sense vibrations requires the determination of the cellular mechanical environment. Here, we quantified vibration-induced fluid shear stresses in vitro and tested whether this system allows for the separation of two mechanical parameters previously proposed to drive the cellular response to vibration – fluid shear and peak accelerations. When peak accelerations of the oscillatory horizontal motions were set at 1g and 60Hz, peak fluid shear stresses acting on the cell layer reached 0.5Pa. A 3.5-fold increase in fluid viscosity increased peak fluid shear stresses 2.6-fold while doubling fluid volume in the well caused a 2-fold decrease in fluid shear. Fluid shear was positively related to peak acceleration magnitude and inversely related to vibration frequency. These data demonstrated that peak shear stress can be effectively separated from peak acceleration by controlling specific levels of vibration frequency, acceleration, and/or fluid viscosity. As an example for exploiting these relations, we tested the relevance of shear stress in promoting COX-2 expression in osteoblast like cells. Across different vibration frequencies and fluid viscosities, neither the level of generated fluid shear nor the frequency of the signal were able to consistently account for differences in the relative increase in COX-2 expression between groups, emphasizing that the eventual identification of the physical mechanism(s) requires a detailed quantification of the cellular mechanical environment. PMID:23074384

Senescence is a cellular response to damage and stress. The senescence response prevents cancer by suppressing the proliferation of cells with a compromised genome and contributes to optimal wound healing in normal tissues. Persistent senescent cells are also thought to drive aging and age-associated pathologies through their secretion of inflammatory factors that modify the tissue microenvironment and alter the function of nearby normal or transformed cells. Understanding how senescent cells alter the microenvironment would be aided by the ability to induce or eliminate senescent cells at will in vivo. Here, we combine the use of the synthetic nucleoside analog ganciclovir (GCV) with herpes simplex virus thymidine kinase (HSVtk) activity to create or eliminate senescent human cells. We show that low concentrations of GCV induce senescence through the accumulation of nuclear DNA damage while higher concentrations of GCV, similar to those used in vivo, kill non-dividing senescent cells via mitochondrial DNA (mtDNA) damage and caspase-dependent apoptosis. Using this system, we effectively eliminated xenografted normal human senescent fibroblasts or induced senescence in human breast cancer cells in vivo. Thus, cellularsenescence and mtDNA damage are outcomes of synthetic nucleoside analog treatment, indicating that the GCV–HSVtk combination can be used effectively to promote the targeted formation or eradication of senescent cells. PMID:23868060

Senescence is a cellular response to damage and stress. The senescence response prevents cancer by suppressing the proliferation of cells with a compromised genome and contributes to optimal wound healing in normal tissues. Persistent senescent cells are also thought to drive aging and age-associated pathologies through their secretion of inflammatory factors that modify the tissue microenvironment and alter the function of nearby normal or transformed cells. Understanding how senescent cells alter the microenvironment would be aided by the ability to induce or eliminate senescent cells at will in vivo. Here, we combine the use of the synthetic nucleoside analog ganciclovir (GCV) with herpes simplex virus thymidine kinase (HSVtk) activity to create or eliminate senescent human cells. We show that low concentrations of GCV induce senescence through the accumulation of nuclear DNA damage while higher concentrations of GCV, similar to those used in vivo, kill non-dividing senescent cells via mitochondrial DNA (mtDNA) damage and caspase-dependent apoptosis. Using this system, we effectively eliminated xenografted normal human senescent fibroblasts or induced senescence in human breast cancer cells in vivo. Thus, cellularsenescence and mtDNA damage are outcomes of synthetic nucleoside analog treatment, indicating that the GCV-HSVtk combination can be used effectively to promote the targeted formation or eradication of senescent cells.

Leaf senescence is a complicated developmental process that involves degenerative changes and nutrient recycling. The progress of leaf senescence is controlled by various environmental cues and plant hormones, including ethylene, jasmonic acid, salicylic acid, abscisic acid, cytokinins, and strigolactones. The production of strigolactones is induced in response to nitrogen and phosphorous deficiency. Strigolactones also accelerate leaf senescence and regulate shoot branching and root architecture. Leaf senescence is actively promoted in a nutrient-poor soil environment, and nutrients are transported from old leaves to young tissues and seeds. Strigolactones might act as important signals in response to nutrient levels in the rhizosphere. In this review, we discuss the possible roles of strigolactones during leaf senescence.

Based on simulations with cellular automaton (CA) traffic flow models, a generic physical feature of the three-phase models studied in the paper is disclosed. The generic feature is a discontinuous character of driver over-acceleration caused by a combination of two qualitatively different mechanisms of over-acceleration: (i) Over-acceleration through lane changing to a faster lane, (ii) over-acceleration occurring in car-following without lane changing. Based on this generic feature a new three-phase CA traffic flow model is developed. This CA model explains the set of the fundamental empirical features of traffic breakdown in real heterogeneous traffic flow consisting of passenger vehicles and trucks. The model simulates also quantitative traffic pattern characteristics as measured in real heterogeneous flow.

Researcher Dr. Yi Li developed a technique to manipulate certain characteristics of plant growth such as anit-senescence. For example, the tobacco leaf was clipped from a transgenic plant (right), and a wildtype plant (left). During ground-based laboratory studies, both leaves were left in a darkened area for 4 months. When retrieved, the wildtype plant leaf was dried-out and the transgenic leaf remained fresh and green. A variation of this technology that involves manipulating plant hormones has been conducted in space-based studies on tomato plants through BioServe Space Technologies. The transport and distribution of auxin, an important plant hormone has shown to be influenced by microgravity, which could lead to improving the quality of fruits and vegetables grown on Earth.

Mammalian cells mostly rely on extracellular molecules to transfer signals to other cells. However, in stress conditions, more robust mechanisms might be necessary to facilitate cell–cell communications. Cellularsenescence, a stress response associated with permanent exit from the cell cycle and the development of an immunogenic phenotype, limits both tumorigenesis and tissue damage. Paradoxically, the long-term presence of senescent cells can promote tissue damage and aging within their microenvironment. Soluble factors secreted from senescent cells mediate some of these cell-nonautonomous effects. However, it is unknown whether senescent cells impact neighboring cells by other mechanisms. Here we show that senescent cells directly transfer proteins to neighboring cells and that this process facilitates immune surveillance of senescent cells by natural killer (NK) cells. We found that transfer of proteins to NK and T cells is increased in the murine preneoplastic pancreas, a site where senescent cells are present in vivo. Proteomic analysis and functional studies of the transferred proteins revealed that the transfer is strictly dependent on cell–cell contact and CDC42-regulated actin polymerization and is mediated at least partially by cytoplasmic bridges. These findings reveal a novel mode of intercellular communication by which senescent cells regulate their immune surveillance and might impact tumorigenesis and tissue aging. PMID:25854920

Senescent cells accumulate during ageing in various tissues and contribute to organismal ageing. However, factors that are involved in the induction of senescence in vivo are still not well understood. SNEV(P) (rp19/) (PSO) (4) is a multifaceted protein, known to be involved in DNA damage repair and senescence, albeit only in vitro. In this study, we used heterozygous SNEV(+/-) mice (SNEV-knockout results in early embryonic lethality) and wild-type littermate controls as a model to elucidate the role of SNEV(P) (rp19/) (PSO) (4) in DNA damage repair and senescence in vivo. We performed PUVA treatment as model system for potently inducing cellularsenescence, consisting of 8-methoxypsoralen in combination with UVA on mouse skin to induce DNA damage and premature skin ageing. We show that SNEV(P) (rp19/) (PSO) (4) expression decreases during organismal ageing, while p16, a marker of ageing in vivo, increases. In response to PUVA treatment, we observed in the skin of both SNEV(P) (rp19/) (PSO) (4) and wild-type mice an increase in γ-H2AX levels, a DNA damage marker. In old SNEV(P) (rp19/) (PSO) (4) mice, this increase is accompanied by reduced epidermis thickening and increase in p16 and collagenase levels. Thus, the DNA damage response occurring in the mouse skin upon PUVA treatment is dependent on SNEV(P) (rp19/) (PSO) (4) expression and lower levels of SNEV(P) (rp19/) (PSO) (4) , as in old SNEV(+/-) mice, result in increase in cellularsenescence and acceleration of premature skin ageing.

Cellularsenescence is a tumor suppressor mechanism where cells enter a permanent growth arrest following cellular stress. Oncogene-induced senescence (OIS) is induced in non-malignant cells following the expression of an oncogene or inactivation of a tumor suppressor. Previously, we have shown that protein kinase C iota (PKCι) depletion induces cellularsenescence in glioblastoma cells in the absence of a detectable DNA damage response. Here we demonstrate that senescent glioblastoma cells exhibit an aberrant centrosome morphology. This was observed in basal levels of senescence, in p21-induced senescence, and in PKCι depletion-induced senescence. In addition, senescent glioblastoma cells are polyploid, Ki-67 negative and arrest at the G1/S checkpoint, as determined by expression of cell cycle regulatory proteins. These markers are all consistent with cells that have undergone mitotic slippage. Failure of the spindle assembly checkpoint to function properly can lead to mitotic slippage, resulting in the premature exit of mitotic cells into the G1 phase of the cell cycle. Although in G1, these cells have the replicated DNA and centrosomal phenotype of a cell that has entered mitosis and failed to divide. Overall, we demonstrate that PKCι depletion initiates mitotic slippage-induced senescence in glioblastoma cells. To our knowledge, this is the first evidence of markers of mitotic slippage directly in senescent cells by co-staining for senescence-associated β-galactosidase and immunofluorescence markers in the same cell population. We suggest that markers of mitotic slippage be assessed in future studies of senescence to determine the extent of mitotic slippage in the induction of cellularsenescence. PMID:26208522

Cellularsenescence is a tumor suppressor mechanism where cells enter a permanent growth arrest following cellular stress. Oncogene-induced senescence (OIS) is induced in non-malignant cells following the expression of an oncogene or inactivation of a tumor suppressor. Previously, we have shown that protein kinase C iota (PKCι) depletion induces cellularsenescence in glioblastoma cells in the absence of a detectable DNA damage response. Here we demonstrate that senescent glioblastoma cells exhibit an aberrant centrosome morphology. This was observed in basal levels of senescence, in p21-induced senescence, and in PKCι depletion-induced senescence. In addition, senescent glioblastoma cells are polyploid, Ki-67 negative and arrest at the G1/S checkpoint, as determined by expression of cell cycle regulatory proteins. These markers are all consistent with cells that have undergone mitotic slippage. Failure of the spindle assembly checkpoint to function properly can lead to mitotic slippage, resulting in the premature exit of mitotic cells into the G1 phase of the cell cycle. Although in G1, these cells have the replicated DNA and centrosomal phenotype of a cell that has entered mitosis and failed to divide. Overall, we demonstrate that PKCι depletion initiates mitotic slippage-induced senescence in glioblastoma cells. To our knowledge, this is the first evidence of markers of mitotic slippage directly in senescent cells by co-staining for senescence-associated β-galactosidase and immunofluorescence markers in the same cell population. We suggest that markers of mitotic slippage be assessed in future studies of senescence to determine the extent of mitotic slippage in the induction of cellularsenescence.

Mesenchymal stem cells (MSCs) contribute to tissue repair in vivo and form an attractive cell source for tissue engineering. Their regenerative potential is impaired by cellularsenescence. The effects of oxidative stress on MSCs are still unknown. Our studies were to investigate into the proliferation potential, cytological features and the telomere linked stress response system of MSCs, subject to acute or prolonged oxidant challenge with hydrogen peroxide. Telomere length was measured using the telomere restriction fragment assay, gene expression was determined by rtPCR. Sub-lethal doses of oxidative stress reduced proliferation rates and induced senescent-morphological features and senescence-associated {beta}-galactosidase positivity. Prolonged low dose treatment with hydrogen peroxide had no effects on cell proliferation or morphology. Sub-lethal and prolonged low doses of oxidative stress considerably accelerated telomere attrition. Following acute oxidant insult p21 was up-regulated prior to returning to initial levels. TRF1 was significantly reduced, TRF2 showed a slight up-regulation. SIRT1 and XRCC5 were up-regulated after oxidant insult and expression levels increased in aging cells. Compared to fibroblasts and chondrocytes, MSCs showed an increased tolerance to oxidative stress regarding proliferation, telomere biology and gene expression with an impaired stress tolerance in aged cells.

Degreening, caused by chlorophyll degradation, is the most obvious symptom of senescing leaves. Chlorophyll degradation can be triggered by endogenous and environmental cues, and ethylene is one of the major inducers. ETHYLENE INSENSITIVE3 (EIN3) is a key transcription factor in the ethylene signaling pathway. It was previously reported that EIN3, miR164, and a NAC (NAM, ATAF, and CUC) transcription factor ORE1/NAC2 constitute a regulatory network mediating leaf senescence. However, how this network regulates chlorophyll degradation at molecular level is not yet elucidated. Here we report a feed-forward regulation of chlorophyll degradation that involves EIN3, ORE1, and chlorophyll catabolic genes (CCGs). Gene expression analysis showed that the induction of three major CCGs, NYE1, NYC1 and PAO, by ethylene was largely repressed in ein3 eil1 double mutant. Dual-luciferase assay revealed that EIN3 significantly enhanced the promoter activity of NYE1, NYC1 and PAO in Arabidopsis protoplasts. Furthermore, Electrophoretic mobility shift assay (EMSA) indicated that EIN3 could directly bind to NYE1, NYC1 and PAO promoters. These results reveal that EIN3 functions as a positive regulator of CCG expression during ethylene-mediated chlorophyll degradation. Interestingly, ORE1, a senescence regulator which is a downstream target of EIN3, could also activate the expression of NYE1, NYC1 and PAO by directly binding to their promoters in EMSA and chromatin immunoprecipitation (ChIP) assays. In addition, EIN3 and ORE1 promoted NYE1 and NYC1 transcriptions in an additive manner. These results suggest that ORE1 is also involved in the direct regulation of CCG transcription. Moreover, ORE1 activated the expression of ACS2, a major ethylene biosynthesis gene, and subsequently promoted ethylene production. Collectively, our work reveals that EIN3, ORE1 and CCGs constitute a coherent feed-forward loop involving in the robust regulation of ethylene-mediated chlorophyll degradation

The plasma membrane is an important cellular organ that perceives incoming developmental and environmental signals and integrates these signals into cellular regulatory mechanisms. It also acts as a barrier against unfavorable extracellular factors to maintain cell viability. Despite its importance for cell viability, molecular components determining cell viability and underlying mechanisms are largely unknown. Here, we show that a plasma membrane-localized MtN3 protein SAG29 regulates cell viability under high salinity in Arabidopsis. The SAG29 gene is expressed primarily in senescing plant tissues. It is induced by osmotic stresses via an abscisic acid-dependent pathway. Whereas the SAG29-overexpressing transgenic plants (35S:SAG29) exhibited an acceleratedsenescence and were hypersensitive to salt stress, the SAG29-deficient mutants were less sensitive to high salinity. Consistent with this, the 35S:SAG29 transgenic plants showed reduced cell viability in the roots under normal growth condition. In contrast, cell viability in the SAG29-deficient mutant roots was indistinguishable from that in the roots of control plants. Notably, the mutant roots exhibited enhanced cell viability under high salinity. Our observations indicate that the senescence-associated SAG29 protein is associated with cell viability under high salinity and other osmotic stress conditions. We propose that the SAG29 protein may serve as a molecular link that integrates environmental stress responses into senescing process.

*The aim of this work was to determine the genetic basis of sugar-regulated senescence and to explore the relationship with other traits, including flowering and nitrogen-use efficiency. *Quantitative trait loci (QTLs) for senescence were mapped in the Arabidopsis Bay-0 x Shahdara recombinant-inbred line (RIL) population after growth on glucose-containing medium, which acceleratessenescence. The extent of whole-rosette senescence was determined by imaging the maximum quantum yield of photosystem II (F(v)/F(m)). *A major QTL on the top of chromosome 4 colocalized with FRI, a major determinant of flowering. This QTL interacted epistatically with a QTL on chromosome 5, where the floral repressor FLC localizes. Vernalization acceleratedsenescence in late-flowering lines with functional FRI and FLC alleles. Comparison with previous results using the Bay-0 x Shahdara population showed that rapid rosette senescence on glucose-containing medium was correlated with early flowering and high sugar content in compost-grown plants. In addition, correlation was found between the expression of flowering and senescence-associated genes in Arabidopsis accessions. However, an additional QTL on chromosome 3 was not linked to flowering, but to nitrogen-use efficiency. *The results show that whole-rosette senescence is genetically linked to the vernalization-dependent control of flowering, but is also controlled by flowering-independent pathways.

Pharmacologic inhibition of the DNA signal transducers Chk1 and p38 blocks G2 arrest and sensitizes glioblastoma cells to chemotherapeutic methylating agent-induced cytotoxicity. Because Akt pathway activation has been suggested to also block G2 arrest induced by DNA-damaging agents and because glioma cells frequently have high levels of Akt activation, we examined the contribution of the Akt pathway to methylating agent-induced G2 arrest and toxicity. U87MG human glioma cells containing an inducible Akt expression construct were incubated with inducing agent or vehicle, after which the cells were exposed to temozolomide and assayed for activation of the components of the G2 arrest pathway and survival. Temozolomide-treated control cells activated the DNA damage signal transducers Chk1, Chk2, and p38, leading to Cdc25C and Cdc2 inactivation, prolonged G2 arrest, and loss of clonagenicity by a combination of senescence and mitotic catastrophe. Temozolomide-treated cells induced to overexpress Akt, however, exhibited significantly less drug-induced Cdc25C/Cdc2 inactivation and less G2 arrest. Akt-mediated suppression of G2 arrest was associated not with alterations in Chk1 or p38 activation but rather with suppression of Chk2 activation and reduced recruitment of Chk2 to sites of damage in chromatin. Unlike bypass of the G2 checkpoint induced by pharmacologic inhibitors of Chk1 or p38, however, Akt-induced bypass of G2 arrest suppressed, rather than enhanced, temozolomide-induced senescence and mitotic catastrophe. These results show that whereas Akt activation suppresses temozolomide-induced Chk2 activation and G2 arrest, the overriding effect is protection from temozolomide-induced cytotoxicity. The Akt pathway therefore represents a new target for the sensitization of gliomas to chemotherapeutic methylating agents such as temozolomide.

Defined as stable cell-cycle arrest, cellularsenescence plays an important role in diverse biological processes including tumorigenesis, organismal aging, and embryonic development. Although increasing evidence has documented the metabolic changes in senescent cells, mitochondrial function and its potential contribution to the fate of senescent cells remain largely unknown. Here, using two in vitro models of cellularsenescence induced by doxorubicin treatment and prolonged passaging of neonatal human foreskin fibroblasts, we report that senescent cells exhibited high ROS level and augmented glucose metabolic rate concomitant with both morphological and quantitative changes of mitochondria. Furthermore, mitochondrial membrane potential depolarized at late stage of senescent cells which eventually led to apoptosis. Our study reveals that mitochondrial hyper-function contributes to the implementation of cellularsenescence and we propose a model in which the mitochondrion acts as the key player in promoting fate-determination in senescent cells. PMID:27056883

Cellularsenescence is a stress response mechanism that limits tumorigenesis and tissue damage. Induction of cellularsenescence commonly coincides with an immunogenic phenotype that promotes self-elimination by components of the immune system, thereby facilitating tumor suppression and limiting excess fibrosis during wound repair. The mechanisms by which senescent cells regulate their immune surveillance are not completely understood. Here we show that ligands of an activating Natural Killer (NK) cell receptor (NKG2D), MICA and ULBP2 are consistently up-regulated following induction of replicative senescence, oncogene-induced senescence and DNA damage - induced senescence. MICA and ULBP2 proteins are necessary for efficient NK-mediated cytotoxicity towards senescent fibroblasts. The mechanisms regulating the initial expression of NKG2D ligands in senescent cells are dependent on a DNA damage response, whilst continuous expression of these ligands is regulated by the ERK signaling pathway. In liver fibrosis, the accumulation of senescent activated stellate cells is increased in mice lacking NKG2D receptor leading to increased fibrosis. Overall, our results provide new insights into the mechanisms regulating the expression of immune ligands in senescent cells and reveal the importance of NKG2D receptor-ligand interaction in protecting against liver fibrosis. PMID:26878797

Jasmonic acid and derivatives are oxylipin signaling compounds derived from linolenic acid. Jasmonates accumulate during natural and dark-induced senescence but the increase in these compounds is not essential for the initiation or progression of these senescence processes. Here we report that during natural and dark-induced senescence the increase in jasmonate levels does not trigger jasmonate signaling. Furthermore we provide evidence that jasmonate production might result from membrane turnover during dark-induced senescence. PMID:21057217

Purpose: To determine whether p16, a molecular marker of cellularsenescence, and CD68, a microglial marker, are detectible in optic nerve glioma tissue stored for decades, thus providing potential targets for pharmacologic intervention. Methods: Cases were retrieved from the Armed Forces Institute of Pathology Registry of Ophthalmic Pathology. Clinical information was tabulated. In specimens with sufficient tissue, a tissue microarray was constructed to conduct molecular studies. Results: Ninety-two cases were included: gender distribution was in a ratio of one male to 1.6 females, and age range was 2 months to 50 years (average age, 10.8 years). Neurofibromatosis type 1 was identified in 10 cases (10.8%). The majority presented with decreased vision and exophthalmos. Forty-eight cases were studied by a tissue microarray construction. Glial fibrillary acidic protein, a control for immunoreactivity, was positive in 46 cases (96%). Immunoreactivity for p16 protein was seen in 36 cases (75%) and CD68-positive cells in 34 (71%). Limitations include referral bias, limited clinical information, limited amount of tissue, and extended period of tissue preservation. Conclusions: Optic nerve glioma is a tumor of the visual axis in young individuals, which is generally indolent but with a variable clinical course. Traditional histopathologic techniques have not been reliably predictive of clinical course. This microarray contains tumors with representative demographic, clinical, and histologic characteristics for optic nerve glioma. Immunoreactivity for p16 protein and CD68 is positive in the majority. These findings suggest a possible explanation for the variable clinical course and identify therapeutic targets in the cell senescence and microglial pathways. PMID:25411512

Considering the early phylogenetic appearance of functional xylem and phloem elements and the range of senescent processes expressed onto genetically, it becomes apparent that such processes are inextricably linked to the evolution, development, reproduction, form, and function of higher plants. The importance of these senescent processes to man are patently obvious since, in one form or another, these processes provide major sources of wood, fiber, and fuel, and are involved in seed development and grain and fruit ripening. To many, the results of senescent processes also have esthetic value including, for example, the grandeur of a Sequoia, the blaze of colors across a desert landscape covered in the spring by ephermal flowers, or the rich tones and panoramic splendor of a deciduous forest in autumn. Senescent processes are widespread, but varied in kind and degree, ranging from whole plants to individual tissues and cells. This symposium was organized primarily around cellular and biochemical aspects of senescence. A major emphasis was the view that senescent processes, and those which developmentally lead to senescence, are highly regulated with an underlying genetic component. Individual papers were processed separately for the database.

Recombinant polynucleotides have expression control sequences that have a senescence responsive element and a minimal promoter, and which are operatively linked to a heterologous nucleotide sequence. The molecules are useful for achieving high levels of expression of genes in senescent cells. Methods of inhibiting expression of genes in senescent cells also are provided.

Oilseed rape (Brassica napus L.) is a crop plant characterized by a poor nitrogen (N) use efficiency that is mainly due to low N remobilization efficiency during the sequential leaf senescence of the vegetative stage. As a high leaf N remobilization efficiency was strongly linked to a high remobilization of proteins during leaf senescence of rapeseed, our objective was to identify senescence-associated protease activities implicated in the protein degradation. To reach this goal, leaf senescence processes and protease activities were investigated in a mature leaf becoming senescent in plants subjected to ample or low nitrate supply. The characterization of protease activities was performed by using in vitro analysis of RuBisCO degradation with or without inhibitors of specific protease classes followed by a protease activity profiling using activity-dependent probes. As expected, the mature leaf became senescent regardless of the nitrate treatment, and nitrate limitation enhanced the senescence processes associated with an enhanced degradation of soluble proteins. The characterization of protease activities revealed that: (i) aspartic proteases and the proteasome were active during senescence regardless of nitrate supply, and (ii) the activities of serine proteases and particularly cysteine proteases (Papain-like Cys proteases and vacuolar processing enzymes) increased when protein remobilization associated with senescence was accelerated by nitrate limitation. Short statement: Serine and particularly cysteine proteases (both PLCPs and VPEs) seem to play a crucial role in the efficient protein remobilization when leaf senescence of oilseed rape was accelerated by nitrate limitation.

Genes of the Sprouty family (Spry1–4) are feedback inhibitors of receptor tyrosine kinase (RTK) signaling. As such, they restrain proliferation of many cell types and have been proposed as tumor-suppressor genes. Although their most widely accepted target is the Extracellular-regulated kinases (ERK) pathway, the mechanisms by which Spry proteins inhibit RTK signaling are poorly understood. In the present work, we describe a novel mechanism by which Spry1 restricts proliferation, independently of the ERK pathway. In vivo analysis of thyroid glands from Spry1 knockout mice reveals that Spry1 induces a senescence-associated secretory phenotype via activation of the NFκB pathway. Consistently, thyroids from Spry1 knockout mice are bigger and exhibit decreased markers of senescence including Ki67 labeling and senescence-associated β-galactosidase. Although such ‘escape' from senescence is not sufficient to promote thyroid tumorigenesis in adult mice up to 5 months, the onset of Phosphatase and tensin homolog (Pten)-induced tumor formation is accelerated when Spry1 is concomitantly eliminated. Accordingly, we observe a reduction of SPRY1 levels in human thyroid malignancies when compared with non-tumoral tissue. We propose that Spry1 acts as a sensor of mitogenic activity that not only attenuates RTK signaling but also induces a cellularsenescence response to avoid uncontrolled proliferation. PMID:24270409

Early-life adversity is associated with acceleratedcellular ageing during development and increased inflammation during adulthood. However, human studies can only establish correlation, not causation, and existing experimental animal approaches alter multiple components of early-life adversity simultaneously. We developed a novel hand-rearing paradigm in European starling nestlings (Sturnus vulgaris), in which we separately manipulated nutritional shortfall and begging effort for a period of 10 days. The experimental treatments accelerated erythrocyte telomere attrition and increased DNA damage measured in the juvenile period. For telomere attrition, amount of food and begging effort exerted additive effects. Only the combination of low food amount and high begging effort increased DNA damage. We then measured two markers of inflammation, high-sensitivity C-reactive protein and interleukin-6, when the birds were adults. The experimental treatments affected both inflammatory markers, though the patterns were complex and different for each marker. The effect of the experimental treatments on adult interleukin-6 was partially mediated by increased juvenile DNA damage. Our results show that both nutritional input and begging effort in the nestling period affect cellular ageing and adult inflammation in the starling. However, the pattern of effects is different for different biomarkers measured at different time points. PMID:28094324

Cellularsenescence contributes to organismal development, aging, and diverse pathologies, yet available assays to detect senescent cells remain unsatisfactory. Here, we designed and synthesized a lipophilic, biotin-linked Sudan Black B (SBB) analogue suitable for sensitive and specific, antibody-enhanced detection of lipofuscin-containing senescent cells in any biological material. This new hybrid histo-/immunochemical method is easy to perform, reliable, and universally applicable to assess senescence in biomedicine, from cancer research to gerontology.

Approaches the regulation of aging by perturbing the rate of senescent changes in studying how the modulating agent exerts its effects. Shows that very subtle molecular changes may be involved in the overall regulation of cellular aging. (Author/BL)

The TERT gene encodes for the reverse transcriptase activity of the telomerase complex and mutations in TERT can lead to dysfunctional telomerase activity resulting in diseases such as dyskeratosis congenita (DKC). Here, we describe a novel TERT mutation at position T1129P leading to DKC with progressive bone marrow (BM) failure in homozygous members of a consanguineous family. BM hematopoietic stem cells (HSCs) of an affected family member were 300-fold reduced associated with a significantly impaired colony forming capacity in vitro and impaired repopulation activity in mouse xenografts. Recent data in yeast suggested improved cellular checkpoint controls by mTOR inhibition preventing cells with short telomeres or DNA damage from dividing. To evaluate a potential therapeutic option for the patient, we treated her primary skin fibroblasts and BM HSCs with the mTOR inhibitor rapamycin. This led to prolonged survival and decreased levels of senescence in T1129P mutant fibroblasts. In contrast, the impaired HSC function could not be improved by mTOR inhibition, as colony forming capacity and multilineage engraftment potential in xenotransplanted mice remained severely impaired. Thus, rapamycin treatment did not rescue the compromised stem cell function of TERTT1129P mutant patient HSCs and outlines limitations of a potential DKC therapy based on rapamycin. PMID:26546739

Cellularsenescence acts as a potent barrier to tumorigenesis and contributes to the anti-tumor activity of certain chemotherapeutic agents. Senescent cells undergo a stable cell cycle arrest controlled by RB and p53 and, in addition, display a senescence-associated secretory phenotype (SASP) involving the production of factors that reinforce the senescence arrest, alter the microenvironment, and trigger immune surveillance of the senescent cells. Through a proteomics analysis of senescent chromatin, we identified the nuclear factor-κB (NF-κB) subunit p65 as a major transcription factor that accumulates on chromatin of senescent cells. We found that NF-κB acts as a master regulator of the SASP, influencing the expression of more genes than RB and p53 combined. In cultured fibroblasts, NF-κB suppression causes escape from immune recognition by natural killer (NK) cells and cooperates with p53 inactivation to bypass senescence. In a mouse lymphoma model, NF-κB inhibition bypasses treatment-induced senescence, producing drug resistance, early relapse, and reduced survival. Our results demonstrate that NF-κB controls both cell-autonomous and non-cell-autonomous aspects of the senescence program and identify a tumor-suppressive function of NF-κB that contributes to the outcome of cancer therapy.

Cellularsenescence acts as a potent barrier to tumorigenesis and contributes to the anti-tumor activity of certain chemotherapeutic agents. Senescent cells undergo a stable cell cycle arrest controlled by RB and p53 and, in addition, display a senescence-associated secretory phenotype (SASP) involving the production of factors that reinforce the senescence arrest, alter the microenvironment, and trigger immune surveillance of the senescent cells. Through a proteomics analysis of senescent chromatin, we identified the nuclear factor-κB (NF-κB) subunit p65 as a major transcription factor that accumulates on chromatin of senescent cells. We found that NF-κB acts as a master regulator of the SASP, influencing the expression of more genes than RB and p53 combined. In cultured fibroblasts, NF-κB suppression causes escape from immune recognition by natural killer (NK) cells and cooperates with p53 inactivation to bypass senescence. In a mouse lymphoma model, NF-κB inhibition bypasses treatment-induced senescence, producing drug resistance, early relapse, and reduced survival. Our results demonstrate that NF-κB controls both cell-autonomous and non-cell-autonomous aspects of the senescence program and identify a tumor-suppressive function of NF-κB that contributes to the outcome of cancer therapy. PMID:21979375

Cancer is increasingly perceived as a systems-level, network phenomenon. The major trend of malignant transformation can be described as a two-phase process, where an initial increase of network plasticity is followed by a decrease of plasticity at late stages of tumor development. The fluctuating intensity of stress factors, like hypoxia, inflammation and the either cooperative or hostile interactions of tumor inter-cellular networks, all increase the adaptation potential of cancer cells. This may lead to the bypass of cellularsenescence, and to the development of cancer stem cells. We propose that the central tenet of cancer stem cell definition lies exactly in the indefinability of cancer stem cells. Actual properties of cancer stem cells depend on the individual "stress-history" of the given tumor. Cancer stem cells are characterized by an extremely large evolvability (i.e. a capacity to generate heritable phenotypic variation), which corresponds well with the defining hallmarks of cancer stem cells: the possession of the capacity to self-renew and to repeatedly re-build the heterogeneous lineages of cancer cells that comprise a tumor in new environments. Cancer stem cells represent a cell population, which is adapted to adapt. We argue that the high evolvability of cancer stem cells is helped by their repeated transitions between plastic (proliferative, symmetrically dividing) and rigid (quiescent, asymmetrically dividing, often more invasive) phenotypes having plastic and rigid networks. Thus, cancer stem cells reverse and replay cancer development multiple times. We describe network models potentially explaining cancer stem cell-like behavior. Finally, we propose novel strategies including combination therapies and multi-target drugs to overcome the Nietzschean dilemma of cancer stem cell targeting: "what does not kill me makes me stronger".

Senescence is characterized by permanent cell-cycle arrest despite continued viability and metabolic activity, in conjunction with the secretion of a complex mixture of extracellular proteins and soluble factors known as the senescence-associated secretory phenotype (SASP). Cellularsenescence has been shown to prevent the proliferation of potentially tumorigenic cells, and is thus generally considered a tumor suppressive process. However, some SASP components may act as pro-tumorigenic mediators on premalignant cells in the microenvironment. A limited number of studies indicated that protein kinase C (PKC) has a role in senescence, with different isoforms having opposing effects. It is therefore important to elucidate the functional role of specific PKCs in senescence. Here we show that PKCη, an epithelial specific and anti-apoptotic kinase, promotes senescence induced by oxidative stress and DNA damage. We further demonstrate that PKCη promotes senescence through its ability to upregulate the expression of the cell cycle inhibitors p21Cip1 and p27Kip1 and enhance transcription and secretion of interleukin-6 (IL-6). Moreover, we demonstrate that PKCη creates a positive loop for reinforcing senescence by increasing the transcription of both IL-6 and IL-6 receptor, whereas the expression of IL-8 is specifically suppressed by PKCη. Thus, the presence/absence of PKCη modulates major components of SASP. Furthermore, we show that the human polymorphic variant of PKCη, 374I, that exhibits higher kinase activity in comparison to WT-374V, is also more effective in IL-6 secretion, p21Cip1 expression and the promotion of senescence, further supporting a role for PKCη in senescence. As there is now considerable interest in senescence activation/elimination to control tumor progression, it is first crucial to reveal the molecular regulators of senescence. This will improve our ability to develop new strategies to harness senescence as a potential cancer therapy in the

Cellularsenescence is historically associated with cancer suppression and aging. Recently, the reach of the senescence genetic program has been extended to include the ability of senescent cells to actively participate in tissue remodelling during many physiological processes, including placental biology, embryonic patterning, wound healing, and tissue stress responses caused by cancer therapy. Besides growth arrest, a significant feature of senescent cells is their ability to modify their immediate microenvironment using a senescence-associated (SA) secretome, commonly termed the SA secretory phenotype (SASP). Among others, the SASP contains growth factors, cytokines, and extracellular proteases that modulate the majority of both the beneficial and detrimental microenvironmental phenotypes caused by senescent cells. The SASP is thus becoming an obvious pharmaceutical target to manipulate SA effects. Herein, we review known signalling pathways underlying the SASP, including the DNA damage response (DDR), stress kinases, inflammasome, alarmin, inflammation- and cell survival-related transcription factors, miRNAs, RNA stability, autophagy, chromatin components, and metabolic regulators. We also describe the SASP as a temporally regulated dynamic sub-program of senescence that can be divided into a rapid DDR-associated phase, an early self-amplification phase, and a late "mature" phase, the late phase currently being the most widely studied SASP signature. Finally, we discuss how deciphering the signalling pathways regulating the SASP reveal targets that can be manipulated to harness the SA effects to benefit therapies for cancer and other age-related pathologies.

Cellularsenescence is a process leading to terminal growth arrest with characteristic morphological features. This process is mediated by telomere-dependent, oncogene-induced and ROS-induced pathways, but persistent DNA damage is the most common cause. Senescence arrest is mediated by p16(INK4a)- and p21(Cip1)-dependent pathways both leading to retinoblastoma protein (pRb) activation. p53 plays a relay role between DNA damage sensing and p21(Cip1) activation. pRb arrests the cell cycle by recruiting proliferation genes to facultative heterochromatin for permanent silencing. Replicative senescence that occurs in hepatocytes in culture and in liver cirrhosis is associated with lack of telomerase activity and results in telomere shortening. Hepatocellular carcinoma (HCC) cells display inactivating mutations of p53 and epigenetic silencing of p16(INK4a). Moreover, they re-express telomerase reverse transcriptase required for telomere maintenance. Thus, senescence bypass and cellular immortality is likely to contribute significantly to HCC development. Oncogene-induced senescence in premalignant lesions and reversible immortality of cancer cells including HCC offer new potentials for tumor prevention and treatment.

Ground-level ozone (O3) concentrations are expected to increase over the 21st century, especially in East Asia. However, the impact of O3 has not been directly assessed at the forest level in this region. We performed O3 flux-based risk assessments of carbon sequestration capacity in an old cool temperate deciduous forest, consisting of O3-sensitive Japanese beech (Fagus crenata), and in a warm temperate deciduous and evergreen forest dominated by O3-tolerant Konara oak (Quercus serrata) based on long-term CO2 flux observations. On the basis of a practical approach for a continuous estimation of canopy-level stomatal conductance (Gs), higher phytotoxic ozone dose above a threshold of 0 uptake (POD0) with higher Gs was observed in the beech forest than that in the oak forest. Light-saturated gross primary production, as a measure of carbon sequestration capacity of forest ecosystem, declined earlier in the late growth season with increasing POD0, suggesting an earlier autumn senescence, especially in the O3-sensitive beech forest, but not in the O3-tolerant oak forest.

Ground-level ozone (O3) concentrations are expected to increase over the 21st century, especially in East Asia. However, the impact of O3 has not been directly assessed at the forest level in this region. We performed O3 flux-based risk assessments of carbon sequestration capacity in an old cool temperate deciduous forest, consisting of O3-sensitive Japanese beech (Fagus crenata), and in a warm temperate deciduous and evergreen forest dominated by O3-tolerant Konara oak (Quercus serrata) based on long-term CO2 flux observations. On the basis of a practical approach for a continuous estimation of canopy-level stomatal conductance (Gs), higher phytotoxic ozone dose above a threshold of 0 uptake (POD0) with higher Gs was observed in the beech forest than that in the oak forest. Light-saturated gross primary production, as a measure of carbon sequestration capacity of forest ecosystem, declined earlier in the late growth season with increasing POD0, suggesting an earlier autumn senescence, especially in the O3-sensitive beech forest, but not in the O3-tolerant oak forest. PMID:27601188

Cellularsenescence is a stable proliferation arrest, a potent tumor suppressor mechanism, and a likely contributor to tissue aging. Cellularsenescence involves extensive cellular remodeling, including of chromatin structure. Autophagy and lysosomes are important for recycling of cellular constituents and cell remodeling. Here we show that an autophagy/lysosomal pathway processes chromatin in senescent cells. In senescent cells, lamin A/C-negative, but strongly γ-H2AX-positive and H3K27me3-positive, cytoplasmic chromatin fragments (CCFs) budded off nuclei, and this was associated with lamin B1 down-regulation and the loss of nuclear envelope integrity. In the cytoplasm, CCFs were targeted by the autophagy machinery. Senescent cells exhibited markers of lysosomal-mediated proteolytic processing of histones and were progressively depleted of total histone content in a lysosome-dependent manner. In vivo, depletion of histones correlated with nevus maturation, an established histopathologic parameter associated with proliferation arrest and clinical benignancy. We conclude that senescent cells process their chromatin via an autophagy/lysosomal pathway and that this might contribute to stability of senescence and tumor suppression.

Tangier disease (TD) is characterized by a deficiency of high density lipoprotein (HDL) in plasma and patients with TD have an increased risk for coronary artery disease (CAD). Recently, we reported that fibroblasts from TD exhibited large and flattened morphology, which is often observed in senescent cells. On the other hand, data have accumulated to show the relationship between cellularsenescence and development of atherosclerotic CAD. The aim of the present study was to investigate whether TD fibroblasts exhibited cellularsenescence. The proliferation of TD fibroblasts was gradually decreased at population doubling level (PDL) {approx}10 compared with control cells. TD cells practically ceased proliferation at PDL {approx}30. DNA synthesis was markedly decreased in TD fibroblasts. TD cells exhibited a higher positive rate for senescence-associated {beta}-galactosidase (SA-{beta}-gal), which is one of the biomarkers of cellularsenescence in vitro. These data showed that TD cells reached cellularsenescence at an earlier PDL compared with controls. Although, there was no difference in the telomere length of fibroblasts between TD and controls at the earlier passage (PDL 6), the telomere length of TD cells was shorter than that of controls at the late passage (PDL 25). Taken together, the current study demonstrates that the late-passaged TD fibroblasts showed senescent phenotype in vitro, which might be related to the increased cardiovascular manifestations in TD patients.

Background Liver fibrosis was viewed as a reversible process. The activation of hepatic stellate cells (HSCs) is a key event in the process of liver fibrosis. The induction of senescence of HSCs would accelerate the clearance of the activated HSCs. Previously, we demonstrated that soluble egg antigens (SEA) of Schistosoma japonicum promoted the senescence of HSCs via STAT3/P53/P21 pathway. In this paper, our study was aimed to explore whether there are other signaling pathways in the process of SEA-induced HSCs aging and the underlying effect of SKP2/P27 signal on senescent HSCs. Methodology/Principal findings Human hepatic stellate cell line, LX-2 cells, were cultured and stimulated with SEA. Western blot and cellular immunofluorescence analysis were performed to determine the expression of senescence-associated protein, such as P27, SKP2 and FoxO3a. Besides, RNA interfering was applied to knockdown the expression of related protein. The senescence of HSCs was determined by senescence-associated β-gal staining. We found that SEA increased the expression of P27 protein, whereas it inhibited the expression of SKP2 and FoxO3a. Knockdown of P27 as well as overexpression of SKP2 both suppressed the SEA-induced senescence of HSCs. In addition, the nuclear translocation of FoxO3a from the nucleus to the cytoplasm was induced by SEA stimulation. Conclusions/Significance The present study demonstrates that SEA promotes HSCs senescence through the FoxO3a/SKP2/P27 pathway. PMID:28036393

We investigated the linear energy transfer (LET) dependence of mutation induction on the hypoxanthine-guanine phosphoribosyl transferase (HPRT) locus in normal human fibroblast-like cells irradiated with accelerated neon-ion beams. The cells were irradiated with neon-ion beams at various LETs ranging from 63 to 335 keV/microm. Neon-ion beams were accelerated by the Riken Ring Cyclotron at the Institute of Physical and Chemical Research in Japan. Mutation induction at the HPRT locus was detected to measure 6-thioguanine-resistant clones. The mutation spectrum of the deletion pattern of exons of mutants was analyzed using the multiplex polymerase chain reaction (PCR). The dose-response curves increased steeply up to 0.5 Gy and leveled off or decreased between 0.5 and 1.0 Gy, compared to the response to (137)Cs gamma-rays. The mutation frequency increased up to 105 keV/microm and then there was a downward trend with increasing LET values. The deletion pattern of exons was non-specific. About 75-100% of the mutants produced using LETs ranging from 63 to 335 keV/mum showed all or partial deletions of exons, while among gamma-ray-induced mutants 30% showed no deletions, 30% partial deletions and 40% complete deletions. These results suggested that the dose-response curves of neon-ion-induced mutations were dependent upon LET values, but the deletion pattern of DNA was not.

Graphene oxide (GO) consisting of a carbon monolayer has been widely investigated for tissue engineering platforms because of its unique properties. For this study, we fabricated a GO-applied scaffold and assessed the cellular and tissue behaviors in the scaffold. A preclinical test was conducted to ascertain whether the GO scaffold promoted bone induction in dog tooth extraction sockets. For this study, GO scaffolds were prepared by coating the surface of a collagen sponge scaffold with 0.1 and 1 µg/mL GO dispersion. Scaffolds were characterized using scanning electron microscopy (SEM), physical testing, cell seeding, and rat subcutaneous implant testing. Then a GO scaffold was implanted into a dog tooth extraction socket. Histological observations were made at 2 weeks postsurgery. SEM observations show that GO attached to the surface of collagen scaffold struts. The GO scaffold exhibited an interconnected structure resembling that of control subjects. GO application improved the physical strength, enzyme resistance, and adsorption of calcium and proteins. Cytocompatibility tests showed that GO application significantly increased osteoblastic MC3T3-E1 cell proliferation. In addition, an assessment of rat subcutaneous tissue response revealed that implantation of 1 µg/mL GO scaffold stimulated cellular ingrowth behavior, suggesting that the GO scaffold exhibited good biocompatibility. The tissue ingrowth area and DNA contents of 1 µg/mL GO scaffold were, respectively, approximately 2.5-fold and 1.4-fold greater than those of the control. Particularly, the infiltration of ED2-positive (M2) macrophages and blood vessels were prominent in the GO scaffold. Dog bone-formation tests showed that 1 µg/mL GO scaffold implantation enhanced bone formation. New bone formation following GO scaffold implantation was enhanced fivefold compared to that in control subjects. These results suggest that GO was biocompatible and had high bone-formation capability for the scaffold

Graphene oxide (GO) consisting of a carbon monolayer has been widely investigated for tissue engineering platforms because of its unique properties. For this study, we fabricated a GO-applied scaffold and assessed the cellular and tissue behaviors in the scaffold. A preclinical test was conducted to ascertain whether the GO scaffold promoted bone induction in dog tooth extraction sockets. For this study, GO scaffolds were prepared by coating the surface of a collagen sponge scaffold with 0.1 and 1 µg/mL GO dispersion. Scaffolds were characterized using scanning electron microscopy (SEM), physical testing, cell seeding, and rat subcutaneous implant testing. Then a GO scaffold was implanted into a dog tooth extraction socket. Histological observations were made at 2 weeks postsurgery. SEM observations show that GO attached to the surface of collagen scaffold struts. The GO scaffold exhibited an interconnected structure resembling that of control subjects. GO application improved the physical strength, enzyme resistance, and adsorption of calcium and proteins. Cytocompatibility tests showed that GO application significantly increased osteoblastic MC3T3-E1 cell proliferation. In addition, an assessment of rat subcutaneous tissue response revealed that implantation of 1 µg/mL GO scaffold stimulated cellular ingrowth behavior, suggesting that the GO scaffold exhibited good biocompatibility. The tissue ingrowth area and DNA contents of 1 µg/mL GO scaffold were, respectively, approximately 2.5-fold and 1.4-fold greater than those of the control. Particularly, the infiltration of ED2-positive (M2) macrophages and blood vessels were prominent in the GO scaffold. Dog bone-formation tests showed that 1 µg/mL GO scaffold implantation enhanced bone formation. New bone formation following GO scaffold implantation was enhanced fivefold compared to that in control subjects. These results suggest that GO was biocompatible and had high bone-formation capability for the scaffold

Cultured human myoblasts fail to immortalize following the introduction of telomerase. The availability of an immortalization protocol for normal human myoblasts would allow one to isolate cellular models from various neuromuscular diseases, thus opening the possibility to develop and test novel therapeutic strategies. The parameters limiting the efficacy of myoblast transfer therapy (MTT) could be assessed in such models. Finally, the presence of an unlimited number of cell divisions, and thus the ability to clone cells after experimental manipulations, reduces the risks of insertional mutagenesis by many orders of magnitude. This opportunity for genetic modification provides an approach for creating a universal donor that has been altered to be more therapeutically useful than its normal counterpart. It can be engineered to function under conditions of chronic damage (which are very different than the massive regeneration conditions that recapitulate normal development), and to overcome the biological problems such as cell death and failure to proliferate and migrate that limit current MTT strategies. We describe here the production and characterization of a human myogenic cell line, LHCN-M2, that has overcome replicative aging due to the expression of telomerase and cyclin-dependent kinase 4. We demonstrate that it functions as well as young myoblasts in xenotransplant experiments in immunocompromized mice under conditions of regeneration following muscle damage.

Summary Cellularsenescence has been implicated in tumor suppression, development, and aging and is accompanied by large-scale chromatin rearrangements, forming senescence-associated heterochromatic foci (SAHF). However, how the chromatin is reorganized during SAHF formation is poorly understood. Furthermore, heterochromatin formation in senescence appears to contrast with loss of heterochromatin in Hutchinson-Gilford progeria. We mapped architectural changes in genome organization in cellularsenescence using Hi-C. Unexpectedly, we find a dramatic sequence- and lamin-dependent loss of local interactions in heterochromatin. This change in local connectivity resolves the paradox of opposing chromatin changes in senescence and progeria. In addition, we observe a senescence-specific spatial clustering of heterochromatic regions, suggesting a unique second step required for SAHF formation. Comparison of embryonic stem cells (ESCs), somatic cells, and senescent cells shows a unidirectional loss in local chromatin connectivity, suggesting that senescence is an endpoint of the continuous nuclear remodelling process during differentiation. PMID:25640177

Floral senescence involves an ordered set of events coordinated at the plant, flower, organ and cellular level. This review assesses our current understanding of the input signals, signal transduction and cellular processes that regulate petal senescence and cell death. In many species a visible sign of petal senescence is wilting. This is accompanied by remobilization of nutrients from the flower to the developing ovary or to other parts of the plant. In other species, petals abscise while still turgid. Coordinating signals for floral senescence also vary across species. In some species ethylene acts as a central regulator, in others floral senescence is ethylene insensitive and other growth regulators are implicated. Due to the variability in this coordination and sequence of events across species, identifying suitable models to study petal senescence has been challenging, and the best candidates are reviewed. Transcriptomic studies provide an overview of the MAP kinases and transcription factors that are activated during petal senescence in several species including Arabidopsis. Our understanding of downstream regulators such as autophagy genes and proteases is also improving. This gives us insights into possible signalling cascades that regulate initiation of senescence and coordination of cell death processes. It also identifies the gaps in our knowledge such as the role of microRNAs. Finally future prospects for using all this information from model to non-model species to extend vase life in ornamental species is reviewed.

Cellularsenescence suppresses cancer by preventing the proliferation of damaged cells, but senescent cells can also promote cancer though the pro-inflammatory senescence-associated secretory phenotype (SASP). Simvastatin, an HMG-coA reductase inhibitor, is known to attenuate inflammation and prevent certain cancers. Here, we show that simvastatin decreases the SASP of senescent human fibroblasts by inhibiting protein prenylation, without affecting the senescent growth arrest. The Rho family GTPases Rac1 and Cdc42 were activated in senescent cells, and simvastatin reduced both activities. Further, geranylgeranyl transferase, Rac1 or Cdc42 depletion reduced IL-6 secretion by senescent cells. We also show that simvastatin mitigates the effects of senescent conditioned media on breast cancer cell proliferation and endocrine resistance. Our findings identify a novel activity of simvastatin and mechanism of SASP regulation. They also suggest that senescent cells, which accumulate after radio/chemo therapy, promote endocrine resistance in breast cancer and that simvastatin might suppress this resistance. PMID:26658759

Glioblastoma (GBM) is a lethal brain tumor composed of heterogeneous cellular populations including glioma stem cells (GSCs) and differentiated non-stem glioma cells (NSGCs). While GSCs are involved in tumor initiation and propagation, NSGCs’ role remains elusive. Here, we demonstrate that NSGCs undergo senescence and secrete pro-angiogenic proteins, boosting the GSC-derived tumor formation in vivo. We used a GSC model that maintains stemness in neurospheres, but loses the stemness and differentiates into NSGCs upon serum stimulation. These NSGCs downregulated telomerase, shortened telomeres, and eventually became senescent. The senescent NSGCs released pro-angiogenic proteins, including vascular endothelial growth factors and senescence-associated interleukins, such as IL-6 and IL-8. Conditioned medium from senescent NSGCs promoted proliferation of brain microvascular endothelial cells, and mixed implantation of GSCs and senescent NSGCs into mice enhanced the tumorigenic potential of GSCs. The senescent NSGCs seem to be clinically relevant, because both clinical samples and xenografts of GBM contained tumor cells that expressed the senescence markers. Our data suggest that senescent NSGCs promote malignant progression of GBM in part via paracrine effects of the secreted proteins. PMID:26775840

The present invention provides a biomarker system for the in vivo and in vitro assessment of cell senescence. In the method of the present invention, {beta}-galactosidase activity is utilized as a means by which cell senescence may be assessed either in in vitro cell cultures or in vivo. 1 fig.

The present invention provides a biomarker system for the in vivo and in vitro assessment of cell senescence. In the method of the present invention, {beta}-galactosidase activity is utilized as a means by which cell senescence may be assessed either in vitro cell cultures or in vivo. 1 fig.

Senescence represents the final stage of leaf development but is often induced prematurely following exposure to biotic and abiotic stresses. Leaf senescence is manifested by color change from green to yellow (due to chlorophyll degradation) or to red (due to de novo synthesis of anthocyanins coupled with chlorophyll degradation) and frequently culminates in programmed death of leaves. However, the breakdown of chlorophyll and macromolecules such as proteins and RNAs that occurs during leaf senescence does not necessarily represent a one-way road to death but rather a reversible process whereby senescing leaves can, under certain conditions, re-green and regain their photosynthetic capacity. This phenomenon essentially distinguishes senescence from programmed cell death, leading researchers to hypothesize that changes occurring during senescence might represent a process of trans-differentiation, that is the conversion of one cell type to another. In this review, we highlight attributes common to senescence and dedifferentiation including chromatin structure and activation of transposable elements and provide further support to the notion that senescence is not merely a deterioration process leading to death but rather a unique developmental state resembling dedifferentiation. PMID:27135333

The present invention provides a biomarker system for the in vivo and in vitro assessment of cell senescence. In the method of the present invention, .beta.-galactosidase activity is utilized as a means by which cell senescence may be assessed either in in vitro cell cultures or in vivo.

The present invention provides a biomarker system for the in vivo and in vitro assessment of cell senescence. In the method of the present invention, .beta.-galactosidase activity is utilized as a means by which cell senescence may be assessed either in vitro cell cultures or in vivo.

Normal and cancer cells facing their demise following exposure to radio-chemotherapy can actively participate in choosing their subsequent fate. These programmed cell fate decisions include true cell death (apoptosis-necroptosis) and therapy-induced cellularsenescence (TIS), a permanent "proliferative arrest" commonly portrayed as premature cellular aging. Despite a permanent loss of proliferative potential, senescent cells remain viable and are highly bioactive at the microenvironment level, resulting in a prolonged impact on tissue architecture and functions. Cellularsenescence is primarily documented as a tumor suppression mechanism that prevents cellular transformation. In the context of normal tissues, cellularsenescence also plays important roles in tissue repair, but contributes to age-associated tissue dysfunction when senescent cells accumulate. Theoretically, in multi-step cancer progression models, cancer cells have already bypassed cellularsenescence during their immortalization step (see hallmarks of cancer). It is then perhaps surprising to find that cancer cells often retain the ability to undergo TIS, or premature aging. This occurs because cellularsenescence results from multiple signalling pathways, some retained in cancer cells, aiming to prevent cell cycle progression in damaged cells. Since senescent cancer cells persist after therapy and secrete an array of cytokines and growth factors that can modulate the tumor microenvironment, these cells may have beneficial and detrimental effects regarding immune modulation and survival of remaining proliferation-competent cancer cells. Similarly, while normal cells undergoing senescence are believed to remain indefinitely growth arrested, whether this is true for senescent cancer cells remains unclear, raising the possibility that these cells may represent a reservoir for cancer recurrence after treatment. This review discusses our current knowledge on cancer cell senescence and highlight questions

This work presents an effective implementation of a numerical model for complete eco-hydrological Cellular Automata modeling on Graphical Processing Units (GPU) with OpenCL (Open Computing Language) for heterogeneous computation (i.e., on CPUs and/or GPUs). Different types of parallel implementations were carried out (e.g., use of fast local memory, loop unrolling, etc), showing increasing performance improvements in terms of speedup, adopting also some original optimizations strategies. Moreover, numerical analysis of results (i.e., comparison of CPU and GPU outcomes in terms of rounding errors) have proven to be satisfactory. Experiments were carried out on a workstation with two CPUs (Intel Xeon E5440 at 2.83GHz), one GPU AMD R9 280X and one GPU nVIDIA Tesla K20c. Results have been extremely positive, but further testing should be performed to assess the functionality of the adopted strategies on other complete models and their ability to fruitfully exploit parallel systems resources.

Cellularsenescence suppresses cancer by arresting cells at risk for malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation. Moreover, using cultured mouse or human fibroblasts and non-malignant breast epithelial cells, we show that senescent fibroblasts disrupt epithelial alveolar morphogenesis, functional differentiation, and branching morphogenesis. Further, we identify MMP-3 as the major factor responsible for the effects of senescent fibroblasts on branching morphogenesis. Our findings support the idea that senescent cells contribute to age-related pathology, including cancer, and describe a new property of senescent fibroblasts--the ability to alter epithelial differentiation--that might also explain the loss of tissue function and organization that is a hallmark of aging.

Phosphodiesterases (PDEs), which hydrolyze and inactivate 3', 5'-cyclic adenosine monophosphate (cAMP) and 3', 5'-cyclic guanosine monophosphate (cGMP), play an important role in synaptic plasticity that underlies memory. Recently, several PDE inhibitors were assessed for their possible therapeutic efficacy in treating cognitive disorders. Here, we examined how cilostazol, a selective PDE3 inhibitor, affects brain functions in senescence-accelerated mouse prone 8 (SAMP8), an animal model of age-related cognitive impairment. Long-term administration of cilostazol restored the impaired context-dependent conditioned fear memory of SAMP8 to match that in normal aging control substrain SAMR1. Cilostazol also increased the number of cells containing phosphorylated cAMP-responsive element binding protein (CREB), a downstream component of the cAMP pathway. Finally, cilostazol improves blood-brain barrier (BBB) integrity, demonstrated by reduced extravasation of 2-deoxy-2-(18)F-fluoro-d-glucose and Evans Blue dye in the brains of SAMP8. This improvement in BBB integrity was associated with an increased amount of zona occludens protein 1 (ZO-1) and occludin proteins, components of tight junctions integral to the BBB. The results suggest that long-term administration of cilostazol exerts its beneficial effects on age-related cognitive impairment through a dual mechanism: by enhancing the cAMP system in the brain and by maintaining or improving BBB integrity.

The phosphate transporter PHT4;6 locates to the trans-Golgi compartment, and its impaired activity causes altered intracellular phosphate compartmentation, leading to low cytosolic Pi levels, a blockage of Golgi-related processes such as protein glycosylation and hemicellulose biosynthesis, and a dwarf phenotype. However, it was unclear whether altered Pi homeostasis in pht4;6 mutants causes further cellular problems, typically associated with limited phosphate availability. Here we report that pht4;6 mutants exhibit a markedly increased disposition to induce dark-induced senescence. In control experiments, in which pht4;6 mutants and wild-type plants developed similarly, we confirmed that accelerated dark-induced senescence in mutants is not a ‘pleiotropic’ process associated with the dwarf phenotype. In fact, accelerated dark-induced senescence in pht4;6 mutants correlates strongly with increased levels of toxic NH4 + and higher sensitivity to ammonium, which probably contribute to the inability of pht4;6 mutants to recover from dark treatment. Experiments with modified levels of either salicylic acid (SA) or trans-zeatin (tZ) demonstrate that altered concentrations of these compounds in pht4;6 plants act as major cellular mediators for dark-induced senescence. This conclusion gained further support from the notion that the expression of the pht4;6 gene is, in contrast to genes coding for major phosphate importers, substantially induced by tZ. Taken together, our findings point to a critical function of PHT4;6 to control cellular phosphate levels, in particular the cytosolic Pi availability, required to energize plant primary metabolism for proper plant development. Phosphate and its allocation mediated by PHT4;6 is critical to prevent onset of dark-induced senescence. PMID:27325894

A sharp definition of what a senescent cell is still lacking since we do not have in depth understanding of mechanisms that induce cellularsenescence. In addition, senescent cells are heterogeneous, in that not all of them express the same genes and present the same phenotype. To further clarify the classification of senescent cells, hints may be derived by the study of cellular metabolism, autophagy and proteasome activity. In this scenario, we decided to study these biological features in senescence of Mesenchymal Stromal Cells (MSC). These cells contain a subpopulation of stem cells that are able to differentiate in mesodermal derivatives (adipocytes, chondrocytes, osteocytes). In addition, they can also contribute to the homeostatic maintenance of many organs, hence, their senescence could be very deleterious for human body functions. We induced MSC senescence by oxidative stress, doxorubicin treatment, X-ray irradiation and replicative exhaustion. The first three are considered inducers of acute senescence while extensive proliferation triggers replicative senescence also named as chronic senescence. In all conditions, but replicative and high IR dose senescence, we detected a reduction of the autophagic flux, while proteasome activity was impaired in peroxide-treated and irradiated cells. Differences were observed also in metabolic status. In general, all senescent cells evidenced metabolic inflexibility and prefer to use glucose as energy fuel. Irradiated cells with low dose of X-ray and replicative senescent cells show a residual capacity to use fatty acids and glutamine as alternative fuels, respectively. Our study may be useful to discriminate among different senescent phenotypes.

Leaf senescence is not a passive degenerative process; it represents a process of nutrient relocation, in which materials are salvaged for growth at a later stage or to produce the next generation. Leaf senescence is regulated by various factors, such as darkness, stress, aging, and phytohormones. Strigolactone is a recently identified phytohormone, and it has multiple functions in plant development, including repression of branching. Although strigolactone is implicated in the regulation of leaf senescence, little is known about its molecular mechanism of action. In this study, strigolactone biosynthesis mutant strains of Arabidopsis (Arabidopsis thaliana) showed a delayed senescence phenotype during dark incubation. The strigolactone biosynthesis genes MORE AXIALLY GROWTH3 (MAX3) and MAX4 were drastically induced during dark incubation and treatment with the senescence-promoting phytohormone ethylene, suggesting that strigolactone is synthesized in the leaf during leaf senescence. This hypothesis was confirmed by a grafting experiment using max4 as the stock and Columbia-0 as the scion, in which the leaves from the Columbia-0 scion senesced earlier than max4 stock leaves. Dark incubation induced the synthesis of ethylene independent of strigolactone. Strigolactone biosynthesis mutants showed a delayed senescence phenotype during ethylene treatment in the light. Furthermore, leaf senescence was strongly accelerated by the application of strigolactone in the presence of ethylene and not by strigolactone alone. These observations suggest that strigolactone promotes leaf senescence by enhancing the action of ethylene. Thus, dark-induced senescence is regulated by a two-step mechanism: induction of ethylene synthesis and consequent induction of strigolactone synthesis in the leaf.

Alterations in lipid metabolism and in the lipid composition of cellular membranes are linked to the pathology of numerous diseases including cancer. However, the influence of oncogene expression on cellular lipid profile is currently unknown. In this work we analyzed changes in lipid profiles that are induced in the course of ERBB2-expression mediated premature senescence. As a model system we used MCF-7 breast cancer cells with doxycycline-inducible expression of NeuT, an oncogenic ERBB2 variant. Affymetrix gene array data showed NeuT-induced alterations in the transcription of many enzymes involved in lipid metabolism, several of which (ACSL3, CHPT1, PLD1, LIPG, MGLL, LDL and NPC1) could be confirmed by quantitative realtime PCR. A study of the glycerophospholipid and lyso-glycerophospholipid profiles, obtained by high performance liquid chromatography coupled to Fourier-transform ion cyclotron resonance-mass spectrometry revealed senescence-associated changes in numerous lipid species, including mitochondrial lipids. The most prominent changes were found in PG(34:1), PG(36:1) (increased) and LPE(18:1), PG(40:7) and PI(36:1) (decreased). Statistical analysis revealed a general trend towards shortened phospholipid acyl chains in senescence and a significant trend to more saturated acyl chains in the class of phosphatidylglycerol. Additionally, the cellular cholesterol content was elevated and accumulated in vacuoles in senescent cells. These changes were accompanied by increased membrane fluidity. In mitochondria, loss of membrane potential along with altered intracellular distribution was observed. In conclusion, we present a comprehensive overview of altered cholesterol and glycerophospholipid patterns in senescence, showing that predominantly mitochondrial lipids are affected and lipid species less susceptible to peroxidation are increased.

Research into extreme drought tolerance in resurrection plants using species such as Craterostigma plantagineum, C. wilmsii, Xerophyta humilis, Tortula ruralis, and Sporobolus stapfianus has provided some insight into the desiccation tolerance mechanisms utilized by these plants to allow them to persist under extremely adverse environmental conditions. Some of the mechanisms used to ensure cellular preservation during severe dehydration appear to be peculiar to resurrection plants. Apart from the ability to preserve vital cellular components during drying and rehydration, such mechanisms include the ability to down-regulate growth-related metabolism rapidly in response to changes in water availability, and the ability to inhibit dehydration-induced senescence programs enabling reconstitution of photosynthetic capacity quickly following a rainfall event. Extensive research on the molecular mechanism of leaf senescence in non-resurrection plants has revealed a multi-layered regulatory network operates to control programed cell death pathways. However, very little is known about the molecular mechanisms that resurrection plants employ to avoid undergoing drought-related senescence during the desiccation process. To survive desiccation, dehydration in the perennial resurrection grass S. stapfianus must proceed slowly over a period of 7 days or more. Leaves detached from the plant before 60% relative water content (RWC) is attained are desiccation-sensitive indicating that desiccation tolerance is conferred in vegetative tissue of S. stapfianus when the leaf RWC has declined to 60%. Whilst some older leaves remaining attached to the plant during dehydration will senesce, suggesting dehydration-induced senescence may be influenced by leaf age or the rate of dehydration in individual leaves, the majority of leaves do not senesce. Rather these leaves dehydrate to air-dryness and revive fully following rehydration. Hence it seems likely that there are genes expressed in

Research into extreme drought tolerance in resurrection plants using species such as Craterostigma plantagineum, C. wilmsii, Xerophyta humilis, Tortula ruralis, and Sporobolus stapfianus has provided some insight into the desiccation tolerance mechanisms utilized by these plants to allow them to persist under extremely adverse environmental conditions. Some of the mechanisms used to ensure cellular preservation during severe dehydration appear to be peculiar to resurrection plants. Apart from the ability to preserve vital cellular components during drying and rehydration, such mechanisms include the ability to down-regulate growth-related metabolism rapidly in response to changes in water availability, and the ability to inhibit dehydration-induced senescence programs enabling reconstitution of photosynthetic capacity quickly following a rainfall event. Extensive research on the molecular mechanism of leaf senescence in non-resurrection plants has revealed a multi-layered regulatory network operates to control programed cell death pathways. However, very little is known about the molecular mechanisms that resurrection plants employ to avoid undergoing drought-related senescence during the desiccation process. To survive desiccation, dehydration in the perennial resurrection grass S. stapfianus must proceed slowly over a period of 7 days or more. Leaves detached from the plant before 60% relative water content (RWC) is attained are desiccation-sensitive indicating that desiccation tolerance is conferred in vegetative tissue of S. stapfianus when the leaf RWC has declined to 60%. Whilst some older leaves remaining attached to the plant during dehydration will senesce, suggesting dehydration-induced senescence may be influenced by leaf age or the rate of dehydration in individual leaves, the majority of leaves do not senesce. Rather these leaves dehydrate to air-dryness and revive fully following rehydration. Hence it seems likely that there are genes expressed in

This paper proposes a real-time feature extraction VLSI architecture for high-resolution images based on the accelerated KAZE algorithm. Firstly, a new system architecture is proposed. It increases the system throughput, provides flexibility in image resolution, and offers trade-offs between speed and scaling robustness. The architecture consists of a two-dimensional pipeline array that fully utilizes computational similarities in octaves. Secondly, a substructure (block-serial discrete-time cellular neural network) that can realize a nonlinear filter is proposed. This structure decreases the memory demand through the removal of data dependency. Thirdly, a hardware-friendly descriptor is introduced in order to overcome the hardware design bottleneck through the polar sample pattern; a simplified method to realize rotation invariance is also presented. Finally, the proposed architecture is designed in TSMC 65 nm CMOS technology. The experimental results show a performance of 127 fps in full HD resolution at 200 MHz frequency. The peak performance reaches 181 GOPS and the throughput is double the speed of other state-of-the-art architectures.

Bangle (Zingiber purpureum) is a tropical ginger that is used as a spice in Southeast Asia. Phenylbutenoid dimers isolated from Bangle have exhibited neurotrophic effects in primary cultured rat cortical neurons and PC12 cells. Furthermore, chronic treatment with phenylbutenoid dimers enhances hippocampal neurogenesis in olfactory bulbectomized mice. In this study, we investigated the effects of Bangle extract on behavior and hippocampal neurogenesis in vivo. SAMP8 mice, which are an established model for accelerated aging, with age-related learning and memory impairments, were given a Bangle-containing diet for 1 month, and subsequent behavioral tests and immunohistochemistry for Ki67, a proliferating cell marker, were performed. We found that the Bangle-containing diet improved spatial learning and memory deficits in the Morris water maze and significantly increased the numbers of Ki67-positive cells in the dentate gyrus of the SAMP8 mice. In addition, the Bangle extract exhibited a neurotrophin-like activity as indicated by the induction of neurite sprouting in PC12 cells. Our results suggest that Bangle is beneficial for the prevention of age-related progression of cognitive impairment.

Effect of the long-term treatment with a Dan-Shen (Salviae miltiorrhizae radix) methanol extract (DME) or its major ingredient lithospermate B (LSB) on memory and learning in scenescence-accelerated mouse (SAM) was investigated by means of Morris's water maze task. DME or LSB treatment significantly decreased the escape latency in the P8 strain of SAM (SAMP8), which strain spontaneously develops learning deficits. These results suggest that DME and LSB treatment improved spacial learning. Neurochemical investigations indicated that both drugs did not affect the cholinergic system, but enhanced functions of the glutamate-PKC system in hippocampus, since the increases in [3H]MK-801 ((+)-5-methyl-10,11-dihydro-5H-dibenzo[a,b]-cyclohepten-5,10-imine maleate) binding in cortex and [3H]PDBu (phorbol 12,13-dibutyrate) binding in hippocampus were observed in DME and LSB treatments. Our previous studies indicate the decrease in PDBu binding and NMDA receptor-mediated functions in SAMP8 brain. Thus, it is suggested that 1) Methanol extract of Dan-Shen is effective for prevention of spacial learning deficit caused with aging, 2) lithospermate B is one of the active ingredients, 3) SAMP8 is useful model animal to investigate drugs for anti-aging or anti-dementia.

The term cellularsenescence was introduced more than five decades ago to describe the state of growth arrest observed in aging cells. Since this initial discovery, the phenotypes associated with cellularsenescence have expanded beyond growth arrest to include alterations in cellular metabolism, secreted cytokines, epigenetic regulation and protein expression. Recently, senescence has been shown to play an important role in vivo not only in relation to aging, but also during embryonic development. Thus, cellularsenescence serves different purposes and comprises a wide range of distinct phenotypes across multiple cell types. Whether all cell types, including post-mitotic neurons, are capable of entering into a senescent state remains unclear. In this review we examine recent data that suggest that cellularsenescence plays a role in brain aging and, notably, may not be limited to glia but also neurons. We suggest that there is a high level of similarity between some of the pathological changes that occur in the brain in Alzheimer's and Parkinson's diseases and those phenotypes observed in cellularsenescence, leading us to propose that neurons and glia can exhibit hallmarks of senescence previously documented in peripheral tissues.

Women experience a series of specific transitions in their reproductive function with age. Shortening of the menstrual cycle begins in the mid to late 30s and is regarded as the first sign of reproductive aging. Other early changes include elevation and increased variance of serum FSH levels, increased incidences of oocyte spindle aberrations and aneuploidy, and declining fertility. The goal of this study was to investigate whether the mouse strain senescence-accelerated mouse-prone-8 (SAMP8) is a suitable model for the study of these midlife reproductive aging characteristics. Midlife SAMP8 mice aged 6.5-7.85 months (midlife SAMP8) exhibited shortened estrous cycles compared with SAMP8 mice aged 2-3 months (young SAMP8, P = .0040). Midlife SAMP8 mice had high FSH levels compared with young SAMP8 mice, and mice with a single day of high FSH exhibited statistically elevated FSH throughout the cycle, ranging from 1.8- to 3.6-fold elevation on the days of proestrus, estrus, metestrus, and diestrus (P < .05). Midlife SAMP8 mice displayed more variance in FSH than young SAMP8 mice (P = .01). Midlife SAMP8 ovulated fewer oocytes (P = .0155). SAMP8 oocytes stained with fluorescently labeled antitubulin antibodies and scored in fluorescence microscopy exhibited increased incidence of meiotic spindle aberrations with age, from 2/126 (1.59%) in young SAMP8 to 38/139 (27.3%) in midlife SAMP8 (17.2-fold increase, P < .0001). Finally, SAMP8 exhibited declining fertility from 8.9 pups/litter in young SAMP8 to 3.5 pups/litter in midlife SAMP8 mice (P < .0001). The age at which these changes occur is younger than for most mouse strains, and their simultaneous occurrence within a single strain has not been described previously. We propose that SAMP8 mice are a model of midlife human female reproductive aging.

Somatic cells senesce in culture after a finite number of divisions indefinitely arresting their proliferation. DNA damage and senescence increase the cellular number of centrosomes, the 2 microtubule organizing centers that ensure bipolar mitotic spindles. Centrosomes also provide the basal body from which primary cilia extend to sense and transduce various extracellular signals, notably Hedgehog. Primary cilium formation is facilitated by cellular quiescence a temporary cell cycle exit, but the impact of senescence on cilia is unknown. We found that senescent human fibroblasts have increased frequency and length of primary cilia. Levels of the negative ciliary regulator CP110 were reduced in senescent cells, as were levels of key elements of the Hedgehog pathway. Hedgehog inhibition reduced proliferation in young cells with increased cilium length accompanying cell cycle arrest suggesting a regulatory function for Hedgehog in primary ciliation. Depletion of CP110 in young cell populations increased ciliation frequencies and reduced cell proliferation. These data suggest that primary cilia are potentially novel determinants of the reduced cellular proliferation that initiates senescence.

Plant senescence is a natural phenomenon known for the appearance of beautiful autumn colors and the ripening of cereals in the field. Senescence is a controlled process that plants utilize to remobilize nutrients from source leaves to developing tissues. While during the past decades, molecular components underlying the onset of senescence have been intensively studied, knowledge remains scarce on the age-dependent mechanisms that control the onset of senescence. Recent advances have uncovered transcriptional networks regulating the competence to senesce. Here, gene regulatory networks acting as internal timing mechanisms for the onset of senescence are highlighted, illustrating that early and late leaf developmental phases are highly connected.

Cellularsenescence has been recently linked to the promotion of age-related pathologies, including a decline in regenerative capacity. While such capacity deteriorates with age in mammals, it remains intact in species such as salamanders, which have an extensive repertoire of regeneration and can undergo multiple episodes through their lifespan. Here we show that, surprisingly, there is a significant induction of cellularsenescence during salamander limb regeneration, but that rapid and effective mechanisms of senescent cell clearance operate in normal and regenerating tissues. Furthermore, the number of senescent cells does not increase upon repetitive amputation or ageing, in contrast to mammals. Finally, we identify the macrophage as a critical player in this efficient senescent cell clearance mechanism. We propose that effective immunosurveillance of senescent cells in salamanders supports their ability to undergo regeneration throughout their lifespan.

Cancer and senescence are both complex transformative processes that dramatically alter many features of cell physiology and their interactions with surrounding tissues. Developing the wide range of cellular features characteristic of these conditions requires profound alterations in global gene expression patterns, which can be achieved by suppressing, activating, or uncoupling cellular gene regulatory pathways. Many genes associated with the initiation and development of tumors are regulated at the level of mRNA decay, frequently through the activity of AU-rich mRNA-destabilizing elements (AREs) located in their 3′-untranslated regions. As such, cellular factors that recognize and control the decay of ARE-containing mRNAs can influence tumorigenic or senescent phenotypes mediated by products of these transcripts. In this review, we discuss evidence showing how suppressed expression and/or activity of the ARE-binding protein tristetraprolin (TTP) can contribute to these processes. Next, we outline current findings linking TTP suppression to exacerbation of individual tumorigenic phenotypes, and the roles of specific TTP substrate mRNAs in mediating these effects. Finally, we survey potential mechanisms that cells may employ to suppress TTP expression in cancer, and propose potential diagnostic and therapeutic strategies that may exploit the relationship between TTP expression and tumor progression or senescence. PMID:22387927

Cellularsenescence is a state of stable cell growth arrest. Activation of oncogenes such as RAS in mammalian cells typically triggers cellularsenescence. Oncogene-induced senescence (OIS) is an important tumor suppression mechanism, and suppression of OIS contributes to cell transformation. Oncogenes trigger senescence through a multitude of incompletely understood downstream signaling events that frequently involve protein kinases. To identify target proteins required for RAS-induced senescence, we developed a small-molecule screen in primary human fibroblasts undergoing senescence induced by oncogenic RAS (H-Ras(G12V)). Using a high-content imaging system to monitor two hallmarks of senescence, senescence-associated β-galactosidase activity expression and inhibition of proliferation, we screened a library of known small-molecule kinase inhibitors for those that suppressed OIS. Identified compounds were subsequently validated and confirmed using a third marker of senescence, senescence-associated heterochromatin foci. In summary, we have established a novel high-content screening platform that may be useful for elucidating signaling pathways mediating OIS by targeting critical pathway components.

Light regulates leaf senescence and light deprivation causes large-scale transcriptional reprogramming to dismantle cellular components and remobilize nutrients to sink organs, such as seeds and storage tissue. We recently reported that in Arabidopsis (Arabidopsis thaliana), Phytochrome-Interacting Factor4 (PIF4) and PIF5 promote dark-induced senescence and natural senescence by directly activating the expression of typical senescence-associated genes (SAGs), including ORESARA1 (ORE1) and ETHYLENE INSENSITIVE3 (EIN3). In contrast, phytochrome B (PhyB) inhibits leaf senescence by repressing PIF4 and PIF5 at the post-translational level. Although we found how red light signaling represses leaf senescence in Arabidopsis, it remains unknown whether PhyB and/or PhyA are involved in leaf senescence in rice (Oryza sativa). Here we show that rice phyB knockout mutants (osphyB-1, -2, and -3) exhibited an early senescence phenotype during dark-induced senescence, but an osphyA knockout mutant (osphyA-3) senesced normally. The RT-qPCR analysis revealed that several senescence-associated genes, including OsORE1 and OsEIN3, were significantly up-regulated in osphyB-2 mutants, indicating that OsPhyB also inhibits leaf senescence, like Arabidopsis PhyB. We also found that leaf segments of osphyB-2 senesced faster even under light conditions. Supplementation with nitrogen compounds, such as KNO3 and NH4NO3, rescued the early senescence phenotype of osphyB-2, indicating that starvation is one of the major signaling factors in the OsPhyB-dependent leaf senescence pathway. PMID:27135344

Paramecium tetraurelia cells have a limited clonal life span and die after approximately 200 fissions if they do not undergo the process of autogamy or conjugation. To test the possibility that cellularsenescence of this species is caused by telomere shortening, we analyzed the genomic DNA of the macronucleus during the clonal life span of P. tetraurelia. We found that telomeric DNA sequences were not shortened during the interval of decreased fission rate and cellular death, defined as senescence in these cells. However, the mean size of the macronuclear DNA was markedly decreased during the clonal life span. We present a model that expands upon previous proposals that accumulated DNA damage causes cellularsenescence in P. tetraurelia. Images PMID:8127914

The efficacy of cell therapy using endothelial colony-forming cells (ECFCs) in the treatment of ischemia is limited by the replicative senescence of isolated ECFCs in vitro. Such senescence must therefore be overcome in order for such cell therapies to be clinically applicable. This study aimed to investigate the potential of sulfated polysaccharide fucoidan to rescue ECFCs from cellularsenescence and to improve in vivo vascular repair by ECFCs. Fucoidan-preconditioning of senescent ECFCs was shown by flow cytometry to restore the expression of functional ECFC surface markers (CD34, c-Kit, VEGFR2, and CXCR4) and stimulate the in vitro tube formation capacity of ECFCs. Fucoidan also promoted the expression of cell cycle-associated proteins (cyclin E, Cdk2, cyclin D1, and Cdk4) in senescent ECFCs, significantly reversed cellularsenescence, and increased the proliferation of ECFCs via the FAK, Akt, and ERK signaling pathways. Fucoidan was found to enhance the survival, proliferation, incorporation, and endothelial differentiation of senescent ECFCs transplanted in ischemic tissues in a murine hind limb ischemia model. Moreover, ECFC-induced functional recovery and limb salvage were markedly improved by fucoidan pretreatment of ECFCs. To our knowledge, the findings of our study are the first to demonstrate that fucoidan enhances the neovasculogenic potential of ECFCs by rescuing them from replicative cellularsenescence. Pretreatment of ECFCs with fucoidan may thus provide a novel strategy for the application of senescent stem cells to therapeutic neovascularization.

Alzheimer's disease amyloid beta protein (Abeta) inhibits cellular reduction of the dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Kaneko et al. have previously hypothesized that Abeta works by suppressing mitochondrial succinate dehydrogenase (SDH), but Liu and Schubert have recently demonstrated that Abeta decreases cellular MTT reduction by accelerating the exocytosis of MTT formazan in neuronal cells. To ask which is the case in astrocytes, we compared the effects of Abeta and 3-nitropropionic acid (3-NP), a specific SDH inhibitor, on MTT reduction in cultured rat cortical astrocytes. Treatment with 3-NP (10 mM) decreased cellular activity of MTT reduction, regardless of the time of incubation with MTT. On the other hand. Abeta-induced inhibition of cellular MTT reduction was dependent on the time of incubation with MTT. The cells treated with Abeta (0.1-1000 nM) exhibited normal capacity for MTT reduction at an early stage of incubation ( < 30 min), but ceased to reduce MTT at the late stage (> 1 h). Microscopic examination revealed that Abeta treatment accelerated the appearance of needle-like MTT formazan crystals at the cell surface. These observations support that Abeta accelerates the exocytosis of MTT formazan in astrocytes. In addition to inhibition of MTT reduction, Abeta is known to induce morphological changes in astrocytes. Following addition of Abeta (20 microM), polygonal astrocytes changed into process-bearing stellate cells. To explore a possible linkage between these two effects of Abeta, we tested if astrocyte stellation is induced by agents that mimic the effect of Abeta on MTT reduction. Cholesterol (5 5000 nM) and lysophosphatidic acid (0.2-20 microg/ml) were found to accelerate the exocytosis of MTT formazan in a similar manner to Abeta, but failed to induce astrocyte stellation. Therefore, Abeta-induced inhibition of MTT reduction is unlikely to be directly linked to its effect on astrocyte morphology.

Cellularsenescence suppresses cancer by eliminating potentially oncogenic cells, participates in tissue repair, contributes to cancer therapy, and promotes organismal aging. Numerous activities of senescent cells depend on the aptitude of these cells to secrete myriads of bioactive molecules, a behavior termed the senescence-associated secretory phenotype (SASP). The SASP supports cell-autonomous functions like the senescence-associated growth arrest, and mediates paracrine interactions between senescent cells and their surrounding microenvironment. The biological functions and the regulation of the SASP are beginning to emerge, and current SASP assessment techniques include the analysis of SASP factors at the mRNA level, the direct measurement of factors inside or outside the cell (i.e., secreted), and the detection of SASP-provoked cellular responses. Here, we focus on a simple approach to collect SASP-conditioned media in order to directly measure secreted SASP factors using sandwich enzyme-linked immunosorbent assay. As an example, we discuss the assessment of the major SASP factor interleukin-6 in senescent human fibroblasts. Supplemental notes are provided to easily adapt this procedure to other SASP factors, change cell types, or scale the techniques for different volumes or high-throughput measurements. These techniques should facilitate the discovery of novel functions and regulators of the SASP.

Aging is a universal, intrinsic, and time-dependent biological decay that is linked to intricate cellular processes including cellularsenescence, telomere shortening, stem cell exhaustion, mitochondrial dysfunction, and deregulated metabolism. Cellularsenescence is accepted as one of the core processes of aging at the organism level. Understanding the molecular mechanism underlying senescence could facilitate the development of potential therapeutics for aging and age-related diseases. Recently, the discovery of long non-coding RNAs (lncRNA) provided insights into a novel regulatory layer that can intervene with cellularsenescence. Increasing evidence indicates that targeting lncRNAs may impact on senescence pathways. In this review, we will focus on lncRNAs involved in mechanistic pathways governing cellularsenescence. PMID:27999563

Thirteen major volatiles of the carnation flower fragrance signature have been identified by GC/MS. Of these, ten, hexanal, (2E)-hexenal, 1-hexanol, 2-hexanol, 3-hexen-1-ol, nonanal, benzaldehyde, benzyl alcohol, benzyl benzoate and caryophyllene, were quantified. The steady-state levels of these ten volatiles change independently as the flowers develop and senesce, suggesting that their synthesis is developmentally regulated. In addition, the chemical composition of the fragrance signature in naturally senesced flowers proved to be very different from that for flowers that had been induced to senesce prematurely by treatment with ethylene. Thus, senescence-related changes in carnation floral scent appear not to be directly regulated by ethylene. From cellular fractionation studies, it is evident that all of the volatiles, except 2-hexanol, are present in both membranous and cytosolic compartments, suggesting that their synthesis is membrane-associated and that they subsequently partition into the cytosol in accordance with partition coefficients.

Cellularsenescence is a fundamental mechanism by which cells remain metabolically active yet cease dividing and undergo distinct phenotypic alterations, including upregulation of p16Ink4a, profound secretome changes, telomere shortening, and decondensation of pericentromeric satellite DNA. Because senescent cells accumulate in multiple tissues with aging, these cells and the dysfunctional factors they secrete, termed the senescence-associated secretory phenotype (SASP), are increasingly recognized as promising therapeutic targets to prevent age-related degenerative pathologies, including osteoporosis. However, the cell type(s) within the bone microenvironment that undergoes senescence with aging in vivo has remained poorly understood, largely because previous studies have focused on senescence in cultured cells. Thus in young (age 6 months) and old (age 24 months) mice, we measured senescence and SASP markers in vivo in highly enriched cell populations, all rapidly isolated from bone/marrow without in vitro culture. In both females and males, p16Ink4a expression by real-time quantitative polymerase chain reaction (rt-qPCR) was significantly higher with aging in B cells, T cells, myeloid cells, osteoblast progenitors, osteoblasts, and osteocytes. Further, in vivo quantification of senescence-associated distension of satellites (SADS), ie, large-scale unraveling of pericentromeric satellite DNA, revealed significantly more senescent osteocytes in old compared with young bone cortices (11% versus 2%, p < 0.001). In addition, primary osteocytes from old mice had sixfold more (p < 0.001) telomere dysfunction-induced foci (TIFs) than osteocytes from young mice. Corresponding with the age-associated accumulation of senescent osteocytes was significantly higher expression of multiple SASP markers in osteocytes from old versus young mice, several of which also showed dramatic age-associated upregulation in myeloid cells. These data show that with aging, a subset of cells

Metabolism is closely linked with cellular state and biological processes, but the mechanisms controlling metabolic properties in different contexts remain unclear. Cellularsenescence is an irreversible growth arrest induced by various stresses, which exhibits active secretory and metabolic phenotypes. Here, we show that retinoblastoma protein (RB) plays a critical role in promoting the metabolic flow by activating both glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) in cells that have undergone oncogene-induced senescence (OIS). A combination of real-time metabolic monitoring, and metabolome and gene expression analyses showed that OIS-induced fibroblasts developed an accelerated metabolic flow. The loss of RB downregulated a series of glycolytic genes and simultaneously reduced metabolites produced from the glycolytic pathway, indicating that RB upregulates glycolytic genes in OIS cells. Importantly, both mitochondrial OXPHOS and glycolytic activities were abolished in RB-depleted or downstream glycolytic enzyme-depleted OIS cells, suggesting that RB-mediated glycolytic activation induces a metabolic flux into the OXPHOS pathway. Collectively, our findings reveal that RB essentially functions in metabolic remodeling and the maintenance of the active energy production in OIS cells.

Metabolism is closely linked with cellular state and biological processes, but the mechanisms controlling metabolic properties in different contexts remain unclear. Cellularsenescence is an irreversible growth arrest induced by various stresses, which exhibits active secretory and metabolic phenotypes. Here, we show that retinoblastoma protein (RB) plays a critical role in promoting the metabolic flow by activating both glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) in cells that have undergone oncogene-induced senescence (OIS). A combination of real-time metabolic monitoring, and metabolome and gene expression analyses showed that OIS-induced fibroblasts developed an accelerated metabolic flow. The loss of RB downregulated a series of glycolytic genes and simultaneously reduced metabolites produced from the glycolytic pathway, indicating that RB upregulates glycolytic genes in OIS cells. Importantly, both mitochondrial OXPHOS and glycolytic activities were abolished in RB-depleted or downstream glycolytic enzyme-depleted OIS cells, suggesting that RB-mediated glycolytic activation induces a metabolic flux into the OXPHOS pathway. Collectively, our findings reveal that RB essentially functions in metabolic remodeling and the maintenance of the active energy production in OIS cells. PMID:26009982

Background: Acceleratedcellularsenescence within the nucleus pulposus (NP) region is a common feature of disc degeneration. Our previous work indicated that TNF-α promoted NP cell senescence. Although the intervertebral disc has been reported to be an estrogen-sensitive tissue, it is unclear whether estrogen can inhibit premature senescence of NP cells. Objective: To investigate whether 17beta-estradiol (E2) can attenuate TNF-α-induced premature senescence of NP cells and the potential mechanism behind this regulatory process. Methods: Isolated NP cells and intact intervertebral discs from healthy rats were cultured with or without TNF-α, E2 or their combination. The pan estrogen receptor (ER) antagonist ICI 182780 was used to investigate the role of ER. Direct and indirect indicators including cell proliferation, SA-β-Gal activity, telomerase activity, cell cycle, and the expression of matrix macromolecules (aggrecan and collagen II) and senescence markers (p16 and p53) were used to evaluate the premature senescence of NP cells. Additionally, intracellular reactive oxygen species (ROS) and NF-κB/p65 activity were also detected in the NP cell cultures. Results: In the NP cell cultures, E2 significantly increased cell proliferation potency, telomerase activity and the expression of matrix macromolecules but attenuated SA-β-Gal activity, senescence marker (p53 and p16) expression and G1 cycle arrest in TNF-α-treated NP cells. Furthermore, E2 inhibited ROS generation and phospho-NF-κB/p65 expression in the TNF-α-treated NP cells. However, the ER antagonist ICI 182780 abolished the effects of E2 on TNF-α-treated NP cells. In the disc organ cultures, E2 also significantly increased matrix synthesis, whereas it decreased senescence marker (p53 and p16) expression, which could be abolished by the ER antagonist ICI 182780. Conclusion: The interaction between E2 and ER can attenuate TNF-α-induced premature senescence of rat NP cells through interfering with the

Senescence-associated secretory phenotype (SASP) is characterized by abundant secretion of various proteins in senescent cells and implicated in tumor progression and inflammatory responses. However, the profile of secreted proteins in SASP is different from cell type to cell type, and currently, universal markers for SASP have not been reported. In the present investigation, we show that SASP-responsive alkaline phosphatase (SASP-RAP) serves as a sensitive, general and convenient marker for SASP. Etoposide-treated cells exhibited a senescent phenotype characterized by senile morphology, positive staining for senescence-associated β-galactosidase, growth arrest and induction of p53 and p21(WAF1/CIP1). In SASP-RAP-transfected cells, exposure to etoposide increased secretion of SASP-RAP time-dependently. The kinetics of secretion was closely correlated with that of activation of the p21(WAF1/CIP1) promoter and the p16(INK4a) promoter. The enhanced secretion of SASP-RAP by senescence was also observed in cells treated with other senescence inducers such as trichostatin A, doxorubicin and 4-phenylbutylic acid. The induction of SASP-RAP by senescence was similarly observed in natural replicative senescence. To confirm selectivity of the SASP-RAP response, cells were treated with senescence-related and -unrelated stimuli (IL-1β, LPS, TNF-α and TGF-β), and induction of senescence markers and activity of SASP-RAP were evaluated in parallel. Unlike etoposide, senescence-unrelated stimuli did not induce p53 and p21(WAF1/CIP1), and it was correlated with lack of induction of SASP-RAP. In contrast, senescence-unrelated stimuli up-regulated conventional indicators for SASP, e.g., MMP-3, IL-6 and TIMP, without induction of senescence. SASP-RAP thus serves as a selective, convenient and general marker for detection and monitoring of SASP during cellularsenescence.

The purpose of this study was to determine whether expression of connective tissue growth factor (CTGF) protein in chronic obstructive pulmonary disease (COPD) is consistent in humans and animal models of COPD and to investigate the role of this protein in lung epithelial cells. CTGF in lung epithelial cells of ex-smokers with COPD was compared with ex-smokers without COPD by immunofluorescence. A total of twenty C57Bl/6 mice and sixteen non-human primates (NHPs) were exposed to cigarette smoke (CS) for 4 weeks. Ten mice of these CS-exposed mice and eight of the CS-exposed NHPs were infected with H3N2 influenza A virus (IAV), while the remaining ten mice and eight NHPs were mock-infected with vehicle as control. Both mRNA and protein expression of CTGF in lung epithelial cells of mice and NHPs were determined. The effects of CTGF overexpression on cell proliferation, p16 protein, and senescence-associated β-galactosidase (SA-β-gal) activity were examined in cultured human bronchial epithelial cells (HBECs). In humans, CTGF expression increased with increasing COPD severity. We found that protein expression of CTGF was upregulated in lung epithelial cells in both mice and NHPs exposed to CS and infected with IAV compared to those exposed to CS only. When overexpressed in HBECs, CTGF acceleratedcellularsenescence accompanied by p16 accumulation. Both CTGF and p16 protein expression in lung epithelia are positively associated with the severity of COPD in ex-smokers. These findings show that CTGF is consistently expressed in epithelial cells of COPD lungs. By accelerating lung epithelial senescence, CTGF may block regeneration relative to epithelial cell loss and lead to emphysema.

Senescence-associated genes are up-regulated during plant senescence and many have been implicated in encoding enzymes involved in the metabolism of senescing tissues. Using the differential display technique, we identified a SAG in bean (Phaseolus vulgaris) leaf that was exclusively expressed during senescence and was designated senescence-associated receptor-like kinase (SARK). The deduced SARK polypeptide consists of a signal peptide, a leucine-rich repeat in the extracellular region, a single membrane-spanning domain, and the characteristic serine/threonine protein kinase domain. The mRNA level for SARK increased prior to the loss of chlorophyll and the decrease of chlorophyll a/b-binding protein mRNA. Detached mature bean leaves, which senesce at an accelerated rate compared with leaves on intact plants, showed a similar temporal pattern of SARK message accumulation. Light and cytokinin, which delayed the initiation of leaf senescence, also delayed SARK gene expression; in contrast, darkness and ethylene, which acceleratedsenescence, advanced the initial appearance of the SARK transcript. SARK protein accumulation exhibited a temporal pattern similar to that of its mRNA. A possible role for SARK in the regulation of leaf senescence was considered. PMID:11080306

Fibroblasts have been extensively used as a model to study cellularsenescence. The purpose of this study was to investigate whether the human foreskin fibroblast aging process could be regulated by using the biomaterial chitosan. Fibroblasts cultured on commercial tissue culture polystyrene (TCPS) entered senescence after 55-60 population doublings (PDs), and were accompanied by larger cell shape, higher senescence-associated β-galactosidase (SA β-gal) activity, lower proliferation capacity, and upregulation of senescence-associated molecular markers p21, p53, retinoblastoma (pRB), and p16. Before senescence was reached, PD48 cells were collected from TCPS and seeded on chitosan for three days (PD48-Cd3) to form multicellular spheroids. The protein expression of senescence-associated secretory phenotypes (SASPs) and senescence-associated molecular markers of these cells in PD48-Cd3 spheroids were downregulated significantly. Following chitosan treatment, fibroblasts reseeded on TCPS showed lower SA β-gal activity, increased cellular motility, and a higher proliferation ability of 70-75 PDs. These phenotypic changes were not accompanied by colonies forming in soft agar and a continuous decrease in the senescence-associated proteins p53 and pRB which act as a barrier to tumorigenesis. These results demonstrate that chitosan treatment could delay the induction of senescence which may be useful and safe for future tissue engineering applications.

ARF is an alternative reading frame product of the INK4a/ARF locus, inactivated in numerous human cancers. ARF is a key regulator of cellularsenescence, an irreversible cell growth arrest that suppresses tumor cell growth. It functions by sequestering MDM2 (a p53 E3 ligase) in the nucleolus, thus activating p53. Besides MDM2, ARF has numerous other interacting partners that induce either cellularsenescence or apoptosis in a p53-independent manner. This further complicates the dynamics of the ARF network. Expression of ARF is frequently disrupted in human cancers, mainly due to epigenetic and transcriptional regulation. Vigorous studies on various transcription factors that either positively or negatively regulate ARF transcription have been carried out. However, recent focus on posttranslational modifications, particularly ubiquitination, indicates wider dynamic controls of ARF than previously known. In this review, we discuss the role and dynamic regulation of ARF in senescence and cancer. PMID:27470213

Cellularsenescence is known as an anti-tumor barrier and is characterized by a number of determinants including cell cycle arrest, senescence associated β-galactosidase activity and secretion of pro-inflammatory mediators. Senescent cells are also subjected to enlargement, cytoskeleton-mediated shape changes and organelle alterations. However, the underlying molecular mechanisms responsible for these last changes remain still uncharacterized. Herein, we have identified the Unfolded Protein Response (UPR) as a player controlling some morphological aspects of the senescent phenotype. We show that senescent fibroblasts exhibit ER expansion and mild UPR activation, but conserve an ER stress adaptive capacity similar to that of exponentially growing cells. By genetically invalidating the three UPR sensors in senescent fibroblasts, we demonstrated that ATF6α signaling dictates senescence-associated cell shape modifications. We also show that ER expansion and increased secretion of the pro-inflammatory mediator IL6 were partly reversed by silencing ATF6α in senescent cells. Moreover, ATF6α drives the increase of senescence associated-β-galactosidase activity. Collectively, these findings unveil a novel and central role for ATF6α in the establishment of morphological features of senescence in normal human primary fibroblasts. PMID:27563820

Sirtuin family members with lysine deacetylase activity are known to play an important role in anti-aging and longevity. Cellularsenescence is one of the hallmarks of aging, and downregulation of sirtuin is reported to induce premature senescence. In this study, we investigated the effects of small-molecule sirtuin inhibitors on cellularsenescence. Various small molecules such as tenovin-1 and EX527 were employed for direct sirtuin activity inhibition. U251, SNB-75, and U87MG glioblastoma cells treated with sirtuin inhibitors exhibited phenotypes with nuclear enlargement. Furthermore, treatment of rat primary astrocytes with tenovin-1 also increased the size of the nucleus. The activity of senescence-associated β-galactosidase, a marker of cellularsenescence, was induced by tenovin-1 and EX527 treatment in U87MG glioblastoma cells. Consistent with the senescent phenotype, treatment with tenovin-1 increased p53 expression in U87MG cells. This study demonstrated the senescence-inducing effect of sirtuin inhibitors, which are potentially useful tools for senescence research.

Mitochondria are central regulators of energy homeostasis and may play a pivotal role in mechanisms of cellularsenescence and age-related neurodegenerative and metabolic disorders. However, mitochondrial bioenergetic parameters have not been systematically evaluated under identi...

Objectives Cellularsenescence is a state of irreversible growth arrest induced either by telomere shortening (replicative senescence) or stress. The bronchial epithelial cell is often injured by inhaled toxic substances, such as cigarette smoke. In the present study, we investigated whether exposure to cigarette smoke extract (CSE) induces senescence of bronchial epithelial cells; and Cordyceps sinensis mechanism of inhibition of CSE-induced cellularsenescence. Methods Human bronchial epithelial cells (16HBE cells) cultured in vitro were treated with CSE and/or C. sinensis. p16, p21, and senescence-associated-galactosidase activity were used to detect cellularsenescence with immunofluorescence, quantitative polymerase chain reaction, and Western blotting. Reactive oxygen species (ROS), PI3K/AKT/mTOR and their phosphorylated proteins were examined to testify the activation of signaling pathway by ROS fluorescent staining and Western blotting. Then, inhibitors of ROS and PI3K were used to further confirm the function of this pathway. Results Cellularsenescence was upregulated by CSE treatment, and C. sinensis can decrease CSE-induced cellularsenescence. Activation of ROS/PI3K/AKT/mTOR signaling pathway was enhanced by CSE treatment, and decreased when C. sinensis was added. Blocking ROS/PI3K/AKT/mTOR signaling pathway can attenuate CSE-induced cellularsenescence. Conclusion CSE can induce cellularsenescence in human bronchial epithelial cells, and ROS/PI3K/AKT/mTOR signaling pathway may play an important role in this process. C. sinensis can inhibit the CSE-induced senescence. PMID:27555762

In the presence of exogenous mortality risks, future reproduction by an individual is worth less than present reproduction to its fitness. Senescent aging thus results inevitably from transferring net fertility into younger ages. Some long-lived organisms appear to defy theory, however, presenting negligible senescence (e.g., hydra) and extended lifespans (e.g., Bristlecone Pine). Here, we investigate the possibility that the onset of vitality loss can be delayed indefinitely, even accepting the abundant evidence that reproduction is intrinsically costly to survival. For an environment with constant hazard, we establish that natural selection itself contributes to increasing density-dependent recruitment losses. We then develop a generalized model of accelerating vitality loss for analyzing fitness optima as a tradeoff between compression and spread in the age profile of net fertility. Across a realistic spectrum of senescent age profiles, density regulation of recruitment can trigger runaway selection for ever-reducing senescence. This novel prediction applies without requirement for special life-history characteristics such as indeterminate somatic growth or increasing fecundity with age. The evolution of nonsenescence from senescence is robust to the presence of exogenous adult mortality, which tends instead to increase the age-independent component of vitality loss. We simulate examples of runaway selection leading to negligible senescence and even intrinsic immortality.

The aim of this study was to analyze whether polyamine (PA) metabolism is involved in dark-induced Hordeum vulgare L. ‘Nagrad’ leaf senescence. In the cell, the titer of PAs is relatively constant and is carefully controlled. Senescence-dependent increases in the titer of the free PAs putrescine, spermidine, and spermine occurred when the process was induced, accompanied by the formation of putrescine conjugates. The addition of the anti-senescing agent cytokinin, which delays senescence, to dark-incubated leaves slowed the senescence-dependent PA accumulation. A feature of the senescence process was initial accumulation of PAs at the beginning of the process and their subsequent decrease during the later stages. Indeed, the process was accompanied by both enhanced expression of PA biosynthesis and catabolism genes and an increase in the activity of enzymes involved in the two metabolic pathways. To confirm whether the capacity of the plant to control senescence might be linked to PA, chlorophyll fluorescence parameters, and leaf nitrogen status in senescing barley leaves were measured after PA catabolism inhibition and exogenously applied γ-aminobutyric acid (GABA). The results obtained by blocking putrescine oxidation showed that the senescence process was accelerated. However, when the inhibitor was applied together with GABA, senescence continued without disruption. On the other hand, inhibition of spermidine and spermine oxidation delayed the process. It could be concluded that in dark-induced leaf senescence, the initial accumulation of PAs leads to facilitating their catabolism. Putrescine supports senescence through GABA production and spermidine/spermine supports senescence-dependent degradation processes, is verified by H2O2 generation. PMID:26779231

Aging is manifest in a variety of changes over time, including changes at the cellular level. Cellular aging acts primarily as a tumor suppressor mechanism, but also may enhance cancer development under certain circumstances. One important process of cellular aging is oncogene-induced senescence, which acts as an important anti-cancer mechanism. Cellularsenescence resulting from damage caused by activated oncogenes prevents the growth or potentially neoplastic cells. Moreover, cells that have entered senescence appear to be targets for elimination by the innnate immune system. In another aspect of cellular aging, the absence of telomerase activity in normal tissues results in such cells lacking a telomere maintenance mechanism. One consequence is that in aging there is an increase in cells with shortened telomeres. In the presence of active oncogenes that cause expansion of a neoplastic clone, shortening of telomeres leading to telomere dysfunction prevents the indefinite expansion of the clone because the cells enter crisis. Crisis results from fusions and other defects caused by dysfunctional telomeres and is a terminal state of the neoplastic clone. In this way the absence of telomerase in human cells, while one cause of cellular aging, also acts as an anti-cancer mechanism. PMID:20705476

Human diploid fibroblasts (HDFs) exposed to subcytotoxic concentrations of oxidative or stressful agents, such as hydrogen peroxide, tert-butylhydroperoxide, or ethanol, undergo stress-induced premature senescence (SIPS). This condition is characterized by the appearance of replicative senescence biomarkers such as irreversible growth arrest, increase in senescence-associated β-galactosidase (SA β-gal) activity, altered cell morphology, and overexpression of several senescence-associated genes. Copper is an essential trace element known to accumulate with ageing and to be involved in the pathogenesis of some age-related disorders. Past studies using either yeast or human cellular models of ageing provided evidence in favor of the role of intracellular copper as a longevity modulator. In the present study, copper ability to cause the appearance of senescent features in HDFs was assessed. WI-38 fibroblasts exposed to a subcytotoxic concentration of copper sulfate presented inhibition of cell proliferation, cell enlargement, increased SA β-gal activity, and mRNA overexpression of several senescence-associated genes such as p21, apolipoprotein J (ApoJ), fibronectin, transforming growth factor β-1 (TGF β1), insulin growth factor binding protein 3, and heme oxygenase 1. Western blotting results confirmed enhanced intracellular p21, ApoJ, and TGF β1 in copper-treated cells. Thus, similar to other SIPS-inducing agents, HDF exposure to subcytotoxic concentration of copper results in premature senescence. Further studies will unravel molecular mechanisms and the biological meaning of copper-associated senescence and lead to a better understanding of copper-related disorder establishment and progression.

Introduction hPTTG1 (human pituitary tumor-transforming gene 1) is an oncogene overexpressed in breast cancer and several other types of cancer. Increased hPTTG1 expression has been shown to be associated with poor patient outcomes in breast cancer. Although hPTTG1 overexpression plays important roles in promoting the proliferation, invasion, and metastasis of cancer cells, it also has been suggested to induce cellularsenescence. Deciphering the mechanism by which hPTTG1 overexpression induces these contradictory actions in breast cancer cells is critical to our understanding of the role of hPTTG1 in breast cancer development. Methods MCF-10A and MCF-7 cells were used to identify the mechanism of hPTTG1-induced senescence. The interplay between hPTTG1 overexpression and chemokine C-X-C motif receptor 2 (CXCR2)/p21-dependent senescence in tumor growth and metastasis of MCF-7 cells was investigated by orthotopic transplantation of severe combined immunodeficiency (SCID) mice. Additionally, human invasive ductal carcinoma (IDC) tissue arrays were used to confirm the hPTTG1/CXCR2/p21 axis established in vitro. Results In this study, we investigated the mechanism of hPTTG1-induced senescence as well as its role in breast cancer progression and metastasis. Herein, we showed that hPTTG1 overexpression reinforced senescence through the CXCR2/p21 signaling. Furthermore, hPTTG1 overexpression activated NF-κB signaling to transactivate the expression of interleukin (IL)-8 and growth-regulated oncogene alpha (GROα) to execute CXCR2 signaling in MCF-7 cells. When CXCR2 expression was knocked down in hPTTG1-overexpressing MCF-7 cells, hPTTG1-induced senescence was abrogated by alleviating CXCR2-induced p21 expression. In a mouse model, CXCR2-mediated senescence limited hPTTG1-induced tumor growth and metastasis. Moreover, CXCR2 knockdown in hPTTG1-overexpressing MCF-7 tumors dramatically accelerated tumor growth and metastasis. Our in vitro and in vivo results demonstrated

The effects of high-fat diet ingestion on senescence-induced modulation of contractile responses to phenylephrine (Phe) were determined in aortas of senescence-accelerated (SAMP8) and non-senescent (SAMR1) mice fed (8weeks) a Western-type high-fat diet (WD). Increased levels of senescence-associated β-galactosidase staining were found in aortas of SAMP8 and SAMR1 with WD. In SAMR1, WD did not modify Phe contraction in spite of inducing major changes in the mechanisms of regulation of contractile responses. Although WD increased NAD(P)H-oxidase-derived O2(-) and augmented peroxynitrite formation, we found an increase of inducible NOS (iNOS)-derived NO production which may contribute to maintain Phe contraction in SAMR1 WD. On SAMP8, WD significantly decreased Phe-induced contractions when compared with SAMP8 under normal chow. This response was not dependent on changes of NOS expression, but rather as consequence of increased antioxidant capacity by superoxide dismutase (SOD1). A similar constrictor influence from cyclooxygenase (COX) pathway on Phe responses was found in SAMR1 and SAMP8 ND. However, WD removed that influence on SAMR1, and produced a switch in the balance from a vasoconstrictor to a vasodilator component in SAMP8. These results were associated to the increased COX-2 expression, suggesting that a COX-2-derived vasodilator prostaglandin may contribute to the vascular adaptations after WD intake. Taken together, our data suggest that WD plays a detrimental role in the vasculature of non-senescent mice by increasing pro-inflammatory (iNOS) and pro-oxidative signaling pathways and may contribute to increase vascular senescence. In senescent vessels, however, WD triggers different intrinsic compensatory alterations which include increase of antioxidant activity by SOD1 and vasodilator prostaglandin production via COX-2.

Heat stress-induced leaf senescence is characterized by the loss of chlorophyll from leaf tissues. The objectives of this study were to examine genetic variations in the level of heat-induced leaf senescence in hybrids of colonial (Agrostis capillaris)×creeping bentgrass (Agrostis stolonifera) contrasting in heat tolerance, and determine whether loss of leaf chlorophyll during heat-induced leaf senescence was due to suppressed chlorophyll synthesis and/or accelerated chlorophyll degradation in the cool-season perennial grass species. Plants of two hybrid backcross genotypes ('ColxCB169' and 'ColxCB190') were exposed to heat stress (38/33°C, day/night) for 28 d in growth chambers. The analysis of turf quality, membrane stability, photochemical efficiency, and chlorophyll content demonstrated significant variations in the level of leaf senescence induced by heat stress between the two genotypes, with ColXCB169 exhibiting a lesser degree of decline in chlorophyll content, photochemical efficiency and membrane stability than ColXCB190. The assays of enzymatic activity or gene expression of several major chlorophyll-synthesizing (porphobilinogen deaminase, Mg-chelatase, protochlorophyllide-reductase) and chlorophyll-degrading enzymes (chlorophyllase, pheophytinase, and chlorophyll-degrading peroxidase) indicated heat-induced decline in leaf chlorophyll content was mainly due to accelerated chlorophyll degradation, as manifested by increased gene expression levels of chlorophyllase and pheophytinase, and the activity of pheophytinase (PPH), while chlorophyll-synthesizing genes and enzymatic activities were not differentially altered by heat stress in the two genotypes. The analysis of heat-induced leaf senescence of pph mutants of Arabidopsis further confirmed that PPH could be one enzymes that plays key roles in regulating heat-accelerated chlorophyll degradation. Further research on enzymes responsible in part for the loss of chlorophyll during heat

Cellular stress increases progressively with aging in mammalian tissues. Chronic stress triggers several signaling cascades that can induce a condition called cellularsenescence. Recent studies have demonstrated that senescent cells express a senescence-associated secretory phenotype (SASP). Emerging evidence indicates that the number of cells expressing biomarkers of cellularsenescence increases in tissues with aging, which implies that cellularsenescence is an important player in organismal aging. In the brain, the aging process is associated with degenerative changes, e.g. synaptic loss and white matter atrophy, which lead to progressive cognitive impairment. There is substantial evidence for the presence of oxidative, proteotoxic and metabolic stresses in aging brain. A low-level, chronic inflammatory process is also present in brain during aging. Astrocytes demonstrate age-related changes that resemble those of the SASP: (i) increased level of intermediate glial fibrillary acidic protein and vimentin filaments, (ii) increased expression of several cytokines and (iii) increased accumulation of proteotoxic aggregates. In addition, in vitro stress evokes a typical senescent phenotype in cultured astrocytes and, moreover, isolated astrocytes from aged brain display the proinflammatory phenotype. All of these observations indicate that astrocytes are capable of triggering the SASP and the astrocytes in aging brain display typical characteristics of cellularsenescence. Bearing in mind the many functions of astrocytes, it is evident that the age-related senescence of astrocytes enhances the decline in functional capacity of the brain. We will review the astroglial changes occurring during aging and emphasize that senescent astrocytes can have an important role in age-related neuroinflammation and neuronal degeneration.

Exposure of MCF-7 breast tumor cells or HCT-116 colon carcinoma cells to clinically relevant concentrations of doxorubicin (Adriamycin; Farmitalia Research Laboratories, Milan, Italy) or camptothecin results in both autophagy and senescence. To determine whether autophagy is required for chemotherapy-induced senescence, reactive oxygen generation induced by Adriamycin was suppressed by N-acetyl cysteine and glutathione, and the induction of ataxia telangiectasia mutated, p53, and p21 was modulated pharmacologically and/or genetically. In all cases, autophagy and senescence were collaterally suppressed. The close association between autophagy and senescence indicated by these experiments reflects their collateral regulation via common signaling pathways. The potential relationship between autophagy and senescence was further examined through pharmacologic inhibition of autophagy with chloroquine and 3-methyl-adenine and genetic ablation of the autophagy-related genes ATG5 and ATG7. However, inhibition of autophagy by pharmacological and genetic approaches could not entirely abrogate the senescence response, which was only reduced and/or delayed. Taken together, our findings suggest that autophagy and senescence tend to occur in parallel, and furthermore that autophagy accelerates the development of the senescent phenotype. However, these responses are not inexorably linked or interdependent, as senescence can occur when autophagy is abrogated. PMID:22927544

Exposure of MCF-7 breast tumor cells or HCT-116 colon carcinoma cells to clinically relevant concentrations of doxorubicin (Adriamycin; Farmitalia Research Laboratories, Milan, Italy) or camptothecin results in both autophagy and senescence. To determine whether autophagy is required for chemotherapy-induced senescence, reactive oxygen generation induced by Adriamycin was suppressed by N-acetyl cysteine and glutathione, and the induction of ataxia telangiectasia mutated, p53, and p21 was modulated pharmacologically and/or genetically. In all cases, autophagy and senescence were collaterally suppressed. The close association between autophagy and senescence indicated by these experiments reflects their collateral regulation via common signaling pathways. The potential relationship between autophagy and senescence was further examined through pharmacologic inhibition of autophagy with chloroquine and 3-methyl-adenine and genetic ablation of the autophagy-related genes ATG5 and ATG7. However, inhibition of autophagy by pharmacological and genetic approaches could not entirely abrogate the senescence response, which was only reduced and/or delayed. Taken together, our findings suggest that autophagy and senescence tend to occur in parallel, and furthermore that autophagy accelerates the development of the senescent phenotype. However, these responses are not inexorably linked or interdependent, as senescence can occur when autophagy is abrogated.

Cellularsenescence refers to irreversible growth arrest of primary eukaryotic cells, a process thought to contribute to aging-related degeneration and disease. Deficiency of RecQ helicase RECQL4 leads to Rothmund–Thomson syndrome (RTS), and we have investigated whether senescence is involved using cellular approaches and a mouse model. We first systematically investigated whether depletion of RECQL4 and the other four human RecQ helicases, BLM, WRN, RECQL1 and RECQL5, impacts the proliferative potential of human primary fibroblasts. BLM-, WRN- and RECQL4-depleted cells display increased staining of senescence-associated β-galactosidase (SA-β-gal), higher expression of p16INK4a or/and p21WAF1 and accumulated persistent DNA damage foci. These features were less frequent in RECQL1- and RECQL5-depleted cells. We have mapped the region in RECQL4 that prevents cellularsenescence to its N-terminal region and helicase domain. We further investigated senescence features in an RTS mouse model, Recql4-deficient mice (Recql4HD). Tail fibroblasts from Recql4HD showed increased SA-β-gal staining and increased DNA damage foci. We also identified sparser tail hair and fewer blood cells in Recql4HD mice accompanied with increased senescence in tail hair follicles and in bone marrow cells. In conclusion, dysfunction of RECQL4 increases DNA damage and triggers premature senescence in both human and mouse cells, which may contribute to symptoms in RTS patients. PMID:24832598

Mutations in the tricarboxylic acid (TCA) cycle enzyme fumarate hydratase (FH) are associated with a highly malignant form of renal cancer. We combined analytical chemistry and metabolic computational modelling to investigate the metabolic implications of FH loss in immortalized and primary mouse kidney cells. Here, we show that the accumulation of fumarate caused by the inactivation of FH leads to oxidative stress that is mediated by the formation of succinicGSH, a covalent adduct between fumarate and glutathione. Chronic succination of GSH, caused by the loss of FH, or by exogenous fumarate, leads to persistent oxidative stress and cellularsenescence in vitro and in vivo. Importantly, the ablation of p21, a key mediator of senescence, in Fh1-deficient mice resulted in the transformation of benign renal cysts into a hyperplastic lesion, suggesting that fumarate-induced senescence needs to be bypassed for the initiation of renal cancers.

Senescence is a highly regulated process that limits cellular replication by enforcing a G1 arrest in response to various stimuli. Replicative senescence occurs in response to telomeric DNA erosion, and telomerase expression can offset replicative senescence leading to immortalization of many human cells. Limited data exists regarding changes of microRNA (miRNA) expression during senescence in human cells and no reports correlate telomerase expression with regulation of senescence-related miRNAs. We used miRNA microarrays to provide a detailed account of miRNA profiles for early passage and senescent human foreskin (BJ) fibroblasts as well as early and late passage immortalized fibroblasts (BJ-hTERT) that stably express the human telomerase reverse transcriptase subunit hTERT. Selected miRNAs that were differentially expressed in senescence were assayed for expression in quiescent cells to identify miRNAs that are specifically associated with senescence-associated growth arrest. From this group of senescence-associated miRNAs, we confirmed the ability of miR-143 to induce growth arrest after ectopic expression in young fibroblasts. Remarkably, miR-143 failed to induce growth arrest in BJ-hTERT cells. Importantly, the comparison of late passage immortalized fibroblasts to senescent wild type fibroblasts reveals that miR-146a, a miRNA with a validated role in regulating the senescence associated secretory pathway, is also regulated during extended cell culture independently of senescence. The discovery that miRNA expression is impacted by expression of ectopic hTERT as well as extended passaging in immortalized fibroblasts contributes to a comprehensive understanding of the connections between telomerase expression, senescence and processes of cellular aging.

Some molecular aspects of flower senescence have been reviewed. The isolation, identification and characterization of different genes from various flowers (mainly from petals) associated with senescence have been discussed. The isolated genes were divided into different groups. A large proportion of genes have been found to be upregulated during flower senescence while some genes were also found to be downregulated indicating that there exists a complex interplay between the expression patterns of various genes. The genes involved in petal expansion are found to be upregulated during normal flower development from anthesis to open flower stage, but XTH (Xyloglucan endotransglucosylase hydrolase) is found to be involved in petal expansion as well as abscission. Cysteine proteases or the genes encoding cysteine proteases (assigned a central role in protein degradation) have been identified from various flower systems, but no cysteine protease has been identified from senescing Mirabilis jalapa flowers. In addition to proteases, the genes encoding ubiquitin (exhibiting proteasomal degradation by 26S proteasomes) have also been identified suggesting the two alternate pathways for protein degradation. Genes encoding specific nucleases have also been identified, but they displayed an early increase in transcript abundance before the senescence symptoms become evident and characterize the involvement of PCD during flower senescence. A range of transcription factors are described and their possible role in flower senescence has been discussed. A detailed description of genes involved in ethylene synthesis and the components involved in ethylene signaling have been presented.

The senescence delaying effect of cytokinin is well known, however, the details behind how this process occurs remain unclear. Efforts to improve understanding of this phenomenon have led to the identification in Arabidopsis of specific cytokinin signaling components through which senescence signal responses are regulated. These include the cytokinin receptor (AHK3), the type-B response regulator (ARR2) and the recently identified cytokinin response factor (CRF6). At the mechanistic end of this process, it was found that increased cell-wall invertase activity which occurs in response to cytokinin is both necessary and sufficient for the inhibition of senescence. Yet, a direct link between the signaling and mechanistic steps of a cytokinin regulated senescence process has yet to be demonstrated. This may be in part because the relationship between senescence and primary metabolism implied by the key role of cell-wall invertase is the subject of two apparently opposing bodies of evidence. Here we briefly summarize and propose a model in which cytokinin mediated changes in sink/source relationships leads to delayed senescence which is consistent with existing evidence both for and against sugars as a trigger for developmental senescence. PMID:23656876

In vivo reprogramming is a promising approach for tissue regeneration in response to injury. Several examples of in vivo reprogramming have been reported in a variety of lineages, but some including skeletal muscle have so far proven refractory. Here, we show that acute and chronic injury enables transcription-factor-mediated reprogramming in skeletal muscle. Lineage tracing indicates that this response frequently originates from Pax7+ muscle stem cells. Injury is associated with accumulation of senescent cells, and advanced aging or local irradiation further enhanced in vivo reprogramming, while selective elimination of senescent cells reduced reprogramming efficiency. The effect of senescence appears to be, at least in part, due to the release of interleukin 6 (IL-6), suggesting a potential link with the senescence-associated secretory phenotype. Collectively, our findings highlight a beneficial paracrine effect of injury-induced senescence on cellular plasticity, which will be important for devising strategies for reprogramming-based tissue repair.

Serotonin, which is well known as a pineal hormone in mammals, plays a key role in conditions such as mood, eating disorders, and alcoholism. In plants, although serotonin has been suggested to be involved in several physiological roles, including flowering, morphogenesis, and adaptation to environmental changes, its regulation and functional roles are as yet not characterized at the molecular level. In this study, we found that serotonin is greatly accumulated in rice (Oryza sativa) leaves undergoing senescence induced by either nutrient deprivation or detachment, and its synthesis is closely coupled with transcriptional and enzymatic induction of the tryptophan biosynthetic genes as well as tryptophan decarboxylase (TDC). Transgenic rice plants that overexpressed TDC accumulated higher levels of serotonin than the wild type and showed delayed senescence of rice leaves. However, transgenic rice plants, in which expression of TDC was suppressed through an RNA interference (RNAi) system, produced less serotonin and senesced faster than the wild type, suggesting that serotonin is involved in attenuating leaf senescence. The senescence-retarding activity of serotonin is associated with its high antioxidant activity compared to either tryptophan or chlorogenic acid. Results of TDC overexpression and TDC RNAi plants suggest that TDC plays a rate-limiting role for serotonin accumulation, but the synthesis of serotonin depends on an absolute amount of tryptophan accumulation by the coordinate induction of the tryptophan biosynthetic genes. In addition, immunolocalization analysis revealed that serotonin was abundant in the vascular parenchyma cells, including companion cells and xylem-parenchyma cells, suggestive of its involvement in maintaining the cellular integrity of these cells for facilitating efficient nutrient recycling from senescing leaves to sink tissues during senescence. PMID:19439571

Adults with diabetes mellitus are much more likely to have cardiovascular disease than those without diabetes mellitus. Genetically engineered mouse models have started to provide important insight into the mechanisms whereby diabetes mellitus promotes atherosclerosis. Such models have demonstrated that diabetes mellitus promotes formation of atherosclerotic lesions, progression of lesions into advanced hemorrhaged lesions, and that it prevents lesion regression. The proatherosclerotic effects of diabetes mellitus are driven in part by the altered function of myeloid cells. The protein S100A9 and the receptor for advanced glycation end-products are important modulators of the effect of diabetes mellitus on myelopoiesis, which might promote monocyte accumulation in lesions. Furthermore, myeloid cell expression of the enzyme acyl-CoA synthetase 1 (ACSL1), which converts long-chain fatty acids into their acyl-CoA derivatives, has emerged as causal to diabetes mellitus-induced lesion initiation. The protective effects of myeloid ACSL1-deficiency in diabetic mice, but not in nondiabetic mice, indicate that myeloid cells are activated by diabetes mellitus through mechanisms that play minor roles in the absence of diabetes mellitus. The roles of reactive oxygen species and insulin resistance in diabetes mellitus-accelerated atherosclerosis are also discussed, primarily in relation to endothelial cells. Translational studies addressing whether the mechanisms identified in mouse models are equally important in humans with diabetes mellitus will be paramount.

Elevated levels of reactive oxygen species (ROS) can attack almost all cell components including genomic DNA to induce many types of DNA damage. In this study, we used Saccharomyces cerevisiae with various mutations in a biological network supposed to prevent deleterious effects of endogenous ROS to test the effect of such a network on yeast chronological aging. Our results showed that cells with defects in cellular antioxidation, DNA repair and DNA damage checkpoints displayed a mutation rate higher than that of wild-type strain. Moreover, the chronological life span of most mutants as determined by colony formation was found to be shorter than that of wild-type cells, especially for the mutants defective in DNA replication and DNA damage checkpoints, although the observed cell number was almost the same for wild-type and mutant strains. The mutants were finally found to be more sensitive to SDS and lysing enzyme treatment, and that the degree of sensitivity was correlated with their chronological life span.

Cellularsenescence is a stable cell cycle arrest that limits the proliferation of pre-cancerous cells. Here we demonstrate that scaffold-attachment-factor A (SAFA) and the long noncoding RNA PANDA differentially interact with polycomb repressive complexes (PRC1 and PRC2) and the transcription factor NF-YA to either promote or suppress senescence. In proliferating cells, SAFA and PANDA recruit PRC complexes to repress the transcription of senescence-promoting genes. Conversely, the loss of SAFA-PANDA-PRC interactions allows expression of the senescence programme. Accordingly, we find that depleting either SAFA or PANDA in proliferating cells induces senescence. However, in senescent cells where PANDA sequesters transcription factor NF-YA and limits the expression of NF-YA-E2F-coregulated proliferation-promoting genes, PANDA depletion leads to an exit from senescence. Together, our results demonstrate that PANDA confines cells to their existing proliferative state and that modulating its level of expression can cause entry or exit from senescence.

Cell senescence is an important tumour suppressor mechanism and driver of ageing. Both functions are dependent on the development of the senescent phenotype, which involves an overproduction of pro-inflammatory and pro-oxidant signals. However, the exact mechanisms regulating these phenotypes remain poorly understood. Here, we show the critical role of mitochondria in cellularsenescence. In multiple models of senescence, absence of mitochondria reduced a spectrum of senescence effectors and phenotypes while preserving ATP production via enhanced glycolysis. Global transcriptomic analysis by RNA sequencing revealed that a vast number of senescent-associated changes are dependent on mitochondria, particularly the pro-inflammatory phenotype. Mechanistically, we show that the ATM, Akt and mTORC1 phosphorylation cascade integrates signals from the DNA damage response (DDR) towards PGC-1β-dependent mitochondrial biogenesis, contributing to aROS-mediated activation of the DDR and cell cycle arrest. Finally, we demonstrate that the reduction in mitochondrial content in vivo, by either mTORC1 inhibition or PGC-1β deletion, prevents senescence in the ageing mouse liver. Our results suggest that mitochondria are a candidate target for interventions to reduce the deleterious impact of senescence in ageing tissues.

Osmotic stress induces several senescence-like processes in leaves, such as specific changes in gene expression and yellowing. These processes are dependent on the accumulation of jasmonates and on intact jasmonate signaling. This chapter describes the treatment of Arabidopsis thaliana leaves with sorbitol as an osmotic stress agent and the determination of the elicited phenotypes encompassing chlorophyll loss, degradation of plastidial membrane lipids, and induction of genes regulated by senescence and jasmonate.

This brief overview of premature senescence of dysfunctional endothelial and endothelial progenitor cells provides information on endothelial cell differentiation and specialization, their ontogeny, and controversies related to endothelial stem and progenitor cells. Stressors responsible for the dysfunction of endothelial and endothelial progenitor cells, as well as cellular mechanisms and consequences of endothelial cell dysfunction are presented. Metabolic signatures of dysfunctional endothelial cells and senescence pathways are described. Emerging strategies to rejuvenate endothelial and endothelial progenitor cells conclude the review. PMID:27451101

Several lines of mouse mammary tissue that had been serially transplanted until mitotic senescence was reached were exposed in vivo to plastic implants that slowly released cholera toxin. Gland tissue surrounding the implants displayed new end buds, indicating reinitiation of growth and morphogenesis. The ability of cholera toxin, which elevates intracellular adenosine 3',5'-monophosphate, to temporarily reverse the senescent phenotype suggests that this mitotic dysfunction results not from generalized cellular deterioration but from specific changes in cell regulation.

In addition to floral senescence and longevity, the control of leaf senescence is a major factor determining the quality of several cut flowers, including Lilium, in the commercial market. To better understand the physiological process underlying leaf senescence in this species, we evaluated: (i) endogenous variation in the levels of phytohormones during leaf senescence, (ii) the effects of leaf darkening in senescence and associated changes in phytohormones, and (iii) the effects of spray applications of abscisic acid (ABA) and pyrabactin on leaf senescence. Results showed that while gibberellin 4 (GA(4)) and salicylic acid (SA) contents decreased, that of ABA increased during the progression of leaf senescence. However, dark-induced senescence increased ABA levels, but did not affect GA(4) and SA levels, which appeared to correlate more with changes in air temperature and/or photoperiod than with the induction of leaf senescence. Furthermore, spray applications of pyrabactin delayed the progression of leaf senescence in cut flowers. Thus, we conclude that (i) ABA plays a major role in the regulation of leaf senescence in Lilium, (ii) darkness promotes leaf senescence and increases ABA levels, and (iii) exogenous applications of pyrabactin inhibit leaf senescence in Lilium, therefore suggesting that it acts as an antagonist of ABA in senescing leaves of cut lily flowers.

Mammalian aging is thought to be partially caused by the diminished capacity of stem/precursor cells to undergo self-renewing divisions. Although many cell-cycle regulators are involved in this process, it is unknown to what extent cell senescence, first identified as irreversible growth arrest in vitro, contributes to the aging process. Here, using a serum-induced mouse oligodendrocyte precursor cell (mOPC) senescence model, we identified esophageal cancer-related gene 4 (Ecrg4) as a senescence inducer with implications for the senescence-like state of postmitotic cells in the aging brain. Although mOPCs could proliferate indefinitely when cultured using the appropriate medium (OPC medium), they became senescent in the presence of serum and maintained their senescent phenotype even when the serum was subsequently replaced by OPC medium. We show that Ecrg4 was up-regulated in the senescent OPCs, its overexpression in OPCs induced senescence by accelerating the proteasome-dependent degradation of cyclins D1 and D3, and that its knockdown by a specific short hairpin RNA prevented these phenotypes. We also show that senescent OPCs secreted Ecrg4 and that recombinant Ecrg4 induced OPC senescence in culture. Moreover, increased Ecrg4 expression was observed in the OPCs and neural precursor cells in the aged mouse brain; this was accompanied by the expression of senescence-associated beta-galactosidase activity, indicating the cells' entrance into senescence. These results suggest that Ecrg4 is a factor linking neural-cell senescence and aging.

Human aging is associated with bone loss leading to bone fragility and increased risk of fractures. The cellular and molecular causes of age-related bone loss are current intensive topic of investigation with the aim of identifying new approaches to abolish its negative effects on the skeleton. Age-related osteoblast dysfunction is the main cause of age-related bone loss in both men and women beyond the fifth decade and results from two groups of pathogenic mechanisms: extrinsic mechanisms that are mediated by age-related changes in bone microenvironment including changes in levels of hormones and growth factors, and intrinsic mechanisms caused by the osteoblast cellularsenescence. The aim of this review is to provide a summary of the intrinsic senescence mechanisms affecting osteoblastic functions and how they can be targeted to abolish age-related osteoblastic dysfunction and bone loss associated with aging.

Glioblastoma multiforme is the most common primary tumor of the central nervous system. The drug temozolomide (TMZ) prolongs lifespan in many glioblastoma patients. The sensitivity of glioblastoma cells to TMZ is interfered by many factors, such as the expression of O-6-methylguanine-DNA methyltransferase (MGMT) and activation of AKT signaling. We have recently identified the interaction between netrin-4 (NTN4) and integrin beta-4 (ITGB4), which promotes glioblastoma cell proliferation via activating AKT-mTOR signaling pathway. In the current work we have explored the effect of NTN4/ITGB4 interaction on TMZ induced glioblastoma cell senescence. We report here that the suppression of either ITGB4 or NTN4 in glioblastoma cell lines significantly enhances cellularsenescence. The sensitivity of GBM cells to TMZ was primarily determined by the expression of MGMT. To omit the effect of MGMT, we concentrated on the cell lines devoid of expression of MGMT. NTN4 partially inhibited TMZ induced cell senescence and rescued AKT from dephosphorylation in U251MG cells, a cell line bearing decent levels of ITGB4. However, addition of exogenous NTN4 displayed no significant effect on TMZ induced senescence rescue or AKT activation in U87MG cells, which expressed ITGB4 at low levels. Furthermore, overexpression of ITGB4 combined with exogenous NTN4 significantly attenuated U87MG cell senescence induced by TMZ. These data suggest that NTN4 protects glioblastoma cells from TMZ induced senescence, probably via rescuing TMZ triggered ITGB4 dependent AKT dephosphorylation. This suggests that interfering the interaction between NTN4 and ITGB4 or concomitant use of the inhibitors of the AKT pathway may improve the therapeutic efficiency of TMZ.

In recent years, the artificial cultivation of Morchella mushrooms that belong to Elata Clade, including Morchella elata, has been developed rapidly in China. However, the prominent problem of spawn aging has been frustrating the morel farming. In this paper, aging in M. elata was achieved from 12 to 17 subcultures and lifespan of 1536-2256 h by successive subculturing. The lifespan can be roughly divided into juvenile phase and senescent phase with respect to the mycelia linear growth rate. After a certain period of rapid growth with almost constant rate at the juvenile phase, the isolate entered the senescent phase characterized by slow down of mycelia growth, producing pigments ahead of time and final death of the apical hyphae. The period of the senescent phase was definitely 240-288 h; while that of the juvenile phase was diverse relying on different isolates. Moreover, microscopic study showed that angles between the leading and primary hyphae increased constantly with aging. In senesced hyphal cells of M. elata, the typical characteristics of autophagy (enlargement of vacuoles and existence of organelles sequestrated lysosomes) and apoptosis (condensation of the cytoplasm and nuclear and plasmolysis) were observed. In addition, in the final stage of senescence, the apical hyphae collapsed with the plasma membrane and all the cellular organelles disrupted, indicated a necrotic mode of cell death. Taken together, these data revealed the involvement of autophagy, apoptosis and necrosis in senescence of M. elata. The characterization and molecular mechanism of autophagy, apoptosis and necrosis need further study and the systematic study of morel aging will be beneficial for the healthy development of morel farming.

Purpose of review The purpose of this review is to provide an update on the cardiac stem cell field with an emphasis on aging and to suggest some relevant strategies directed toward rejuvenation of the senescent heart. Recent findings Stem cells were long considered as a fountain of youth and were assumed to be equipped against any form of aging effect. However, it is now clear that stem cells suffer the consequences of aging as well. With the discovery that cardiac stem cells reside in the heart comes the question whether these cells are also impaired upon aging. As cardiac stem cell properties are also altered with age, autologous stem cell-based therapy to treat heart failure will benefit from new improved strategies. Summary With the goal to improve stem cell properties that are impaired upon aging, some strategies are highlighted. Genetic modification of adult human cardiac progenitor cells prior to autologous stem cell-based therapy, delivery of the next generation of stem cells such as CardioChimeras and CardioClusters, and improvement of the myocardial environment with rejuvenating factors constitute some of the possibilities and are discussed in more detail in this review. PMID:25760821

Curcuminoids are well known for their capabilities to combat risk factors that are associated with ageing and cellularsenescence. Recent reports have demonstrated that curcuminoids can extend the lifespan of model organisms. However, the underlying mechanisms by which these polyphenic compounds exert these beneficial effects remain unknown. In this study, t-BHP-induced premature senescence model in human fibroblasts was chosen to explore the protective effects of a curcuminoid, bisdemethoxycurcumin (BDMC), on cellularsenescence. The results demonstrated that BDMC attenuated oxidative stress-induced senescence-like features which include the induction of an enlarged cellular appearance, higher frequency of senescence-associated β -galactosidase staining activity, appearance of senescence-associated heterochromatic foci in nuclei, decrease in proliferation capability, and alteration in related molecules such as p16 and retinoblastoma protein. Notably, we found that BDMC treatment activated Sirt1/AMPK signaling pathway. Moreover, downregulating Sirt1 by the pharmacological inhibitor nicotianamine or small interfering RNA blocked BDMC-mediated protection against t-BHP-mediated decrease in proliferation. These results suggested that BDMC prevented t-BHP-induced cellularsenescence, and BDMC-induced Sirt1 may be a mechanism mediating its beneficial effects.

Cellularsenescence is classified into two groups: replicative and premature senescence. Gene expression and epigenetic changes are reported to differ between these two groups and cell types. Normal human diploid fibroblast TIG-3 cells have often been used in cellularsenescence research; however, their epigenetic profiles are still not fully understood. To elucidate how cellularsenescence is epigenetically regulated in TIG-3 cells, we analyzed the gene expression and DNA methylation profiles of three types of senescent cells, namely, replicatively senescent, ras-induced senescent (RIS), and non-permissive temperature-induced senescent SVts8 cells, using gene expression and DNA methylation microarrays. The expression of genes involved in the cell cycle and immune response was commonly either down- or up-regulated in the three types of senescent cells, respectively. The altered DNA methylation patterns were observed in replicatively senescent cells, but not in prematurely senescent cells. Interestingly, hypomethylated CpG sites detected on non-CpG island regions (“open sea”) were enriched in immune response-related genes that had non-CpG island promoters. The integrated analysis of gene expression and methylation in replicatively senescent cells demonstrated that differentially expressed 867 genes, including cell cycle- and immune response-related genes, were associated with DNA methylation changes in CpG sites close to the transcription start sites (TSSs). Furthermore, several miRNAs regulated in part through DNA methylation were found to affect the expression of their targeted genes. Taken together, these results indicate that the epigenetic changes of DNA methylation regulate the expression of a certain portion of genes and partly contribute to the introduction and establishment of replicative senescence. PMID:28158250

Increased maternal age at reproduction is often associated with decreased offspring performance in numerous species of plants and animals (including humans). Current evolutionary theory considers such maternal effect senescence as part of a unified process of reproductive senescence, which is under identical age-specific selective pressures to fertility. We offer a novel theoretical perspective by combining William Hamilton’s evolutionary model for aging with a quantitative genetic model of indirect genetic effects. We demonstrate that fertility and maternal effect senescence are likely to experience different patterns of age-specific selection and thus can evolve to take divergent forms. Applied to neonatal survival, we find that selection for maternal effects is the product of age-specific fertility and Hamilton’s age-specific force of selection for fertility. Population genetic models show that senescence for these maternal effects can evolve in the absence of reproductive or actuarial senescence; this implies that maternal effect aging is a fundamentally distinct demographic manifestation of the evolution of aging. However, brief periods of increasingly beneficial maternal effects can evolve when fertility increases with age faster than cumulative survival declines. This is most likely to occur early in life. Our integration of theory provides a general framework with which to model, measure, and compare the evolutionary determinants of the social manifestations of aging. Extension of our maternal effects model to other ecological and social contexts could provide important insights into the drivers of the astonishing diversity of lifespans and aging patterns observed among species. PMID:26715745

1. Senescence, the physiological decline that results in decreasing survival and/or reproduction with age, remains one of the most perplexing topics in biology. Most theories explaining the evolution of senescence (i.e. antagonistic pleiotropy, accumulation of mutations, disposable soma) were developed decades ago. Even though these theories have implicitly focused on unitary animals, they have also been used as the foundation from which the universality of senescence across the tree of life is assumed. 2. Surprisingly, little is known about the general patterns, causes and consequences of whole-individual senescence in the plant kingdom. There are important differences between plants and most animals, including modular architecture, the absence of early determination of cell lines between the soma and gametes, and cellular division that does not always shorten telomere length. These characteristics violate the basic assumptions of the classical theories of senescence and therefore call the generality of senescence theories into question. 3. This Special Feature contributes to the field of whole-individual plant senescence with five research articles addressing topics ranging from physiology to demographic modelling and comparative analyses. These articles critically examine the basic assumptions of senescence theories such as age-specific gene action, the evolution of senescence regardless of the organism's architecture and environmental filtering, and the role of abiotic agents on mortality trajectories. 4. Synthesis. Understanding the conditions under which senescence has evolved is of general importance across biology, ecology, evolution, conservation biology, medicine, gerontology, law and social sciences. The question ‘why is senescence universal or why is it not?’ naturally calls for an evolutionary perspective. Senescence is a puzzling phenomenon, and new insights will be gained by uniting methods, theories and observations from formal demography

. Nuclear export is initiated by ROS-induced phosphorylation of tyrosine 707 within hTERT by the Src kinase family. It might be presumed that protection of mitochondria against oxidative stress is an important telomere length-independent function for telomerase in cell survival. Biotechnology companies are focused on development of therapeutic telomerase vaccines, telomerase inhibitors, and telomerase promoter-driven cell killing in oncology, have a telomerase antagonist in late preclinical studies. Anti-aging medicine-oriented groups have intervened on the market with products working on telomerase activation for a broad range of degenerative diseases in which replicative senescence or telomere dysfunction may play an important role. Since oxidative damage has been shown to shorten telomeres in tissue culture models, the adequate topical, transdermal, or systemic administration of antioxidants (such as, patented ocular administration of 1% N-acetylcarnosine lubricant eye drops in the treatment of cataracts) may be beneficial at preserving telomere lengths and delaying the onset or in treatment of disease in susceptible individuals. Therapeutic strategies toward controlled transient activation of telomerase are targeted to cells and replicative potential in cell-based therapies, tissue engineering and regenerative medicine.

Senescence in biofuel grasses is a critical issue because early senescence decreases potential biomass production by limiting aerial growth and development. 2-Dimensional, differential in-gel electrophoresis (2D-DIGE) followed by mass spectrometry of selected protein spots was used to evaluate differences between leaf proteomes of early (ES)- and late- senescing (LS) genotypes of Prairie cordgrass (ES/LS PCG) and switchgrass (ES/LS SG), just before and after senescence was initiated. Analysis of the manually filtered and statistically evaluated data indicated that 69 proteins were significantly differentially abundant across all comparisons, and a majority (41%) were associated with photosynthetic processes as determined by gene ontology analysis. Ten proteins were found in common between PCG and SG, and nine and 18 proteins were unique to PCG and SG respectively. Five of the 10 differentially abundant spots common to both species were increased in abundance, and five were decreased in abundance. Leaf proteomes of the LS genotypes of both grasses analyzed before senescence contained significantly higher abundances of a 14-3-3 like protein and a glutathione-S-transferase protein when compared to the ES genotypes, suggesting differential cellular metabolism in the LS vs. the ES genotypes. The higher abundance of 14-3-3 like proteins may be one factor that impacts the senescence process in both LS PCG and LS SG. Aconitase dehydratase was found in greater abundance in all four genotypes after the onset of senescence, consistent with literature reports from genetic and transcriptomic studies. A Rab protein of the Ras family of G proteins and an s-adenosylmethionine synthase were more abundant in ES PCG when compared with the LS PCG. In contrast, several proteins associated with photosynthesis and carbon assimilation were detected in greater abundance in LS PCG when compared to ES PCG, suggesting that a loss of these proteins potentially contributed to the ES phenotype

Senescence in biofuel grasses is a critical issue because early senescence decreases potential biomass production by limiting aerial growth and development. 2-Dimensional, differential in-gel electrophoresis (2D-DIGE) followed by mass spectrometry of selected protein spots was used to evaluate differences between leaf proteomes of early (ES)- and late- senescing (LS) genotypes of Prairie cordgrass (ES/LS PCG) and switchgrass (ES/LS SG), just before and after senescence was initiated. Analysis of the manually filtered and statistically evaluated data indicated that 69 proteins were significantly differentially abundant across all comparisons, and a majority (41%) were associated with photosynthetic processes as determined by gene ontology analysis. Ten proteins were found in common between PCG and SG, and nine and 18 proteins were unique to PCG and SG respectively. Five of the 10 differentially abundant spots common to both species were increased in abundance, and five were decreased in abundance. Leaf proteomes of the LS genotypes of both grasses analyzed before senescence contained significantly higher abundances of a 14-3-3 like protein and a glutathione-S-transferase protein when compared to the ES genotypes, suggesting differential cellular metabolism in the LS vs. the ES genotypes. The higher abundance of 14-3-3 like proteins may be one factor that impacts the senescence process in both LS PCG and LS SG. Aconitase dehydratase was found in greater abundance in all four genotypes after the onset of senescence, consistent with literature reports from genetic and transcriptomic studies. A Rab protein of the Ras family of G proteins and an s-adenosylmethionine synthase were more abundant in ES PCG when compared with the LS PCG. In contrast, several proteins associated with photosynthesis and carbon assimilation were detected in greater abundance in LS PCG when compared to ES PCG, suggesting that a loss of these proteins potentially contributed to the ES phenotype

The increasing incidence of osteoporosis worldwide requires anabolic treatments that are safe, effective, and, critically, inexpensive given the prevailing overburdened health care systems. While vigorous skeletal loading is anabolic and holds promise, deficits in mechanotransduction accrued with age markedly diminish the efficacy of readily complied, exercise-based strategies to combat osteoporosis in the elderly. Our approach to explore and counteract these age-related deficits was guided by cellular signaling patterns across hierarchical scales and by the insight that cell responses initiated during transient, rare events hold potential to exert high-fidelity control over temporally and spatially distant tissue adaptation. Here, we present an agent-based model of real-time Ca2+/NFAT signaling amongst bone cells that fully described periosteal bone formation induced by a wide variety of loading stimuli in young and aged animals. The model predicted age-related pathway alterations underlying the diminished bone formation at senescence, and hence identified critical deficits that were promising targets for therapy. Based upon model predictions, we implemented an in vivo intervention and show for the first time that supplementing mechanical stimuli with low-dose Cyclosporin A can completely rescue loading induced bone formation in the senescent skeleton. These pre-clinical data provide the rationale to consider this approved pharmaceutical alongside mild physical exercise as an inexpensive, yet potent therapy to augment bone mass in the elderly. Our analyses suggested that real-time cellular signaling strongly influences downstream bone adaptation to mechanical stimuli, and quantification of these otherwise inaccessible, transient events in silico yielded a novel intervention with clinical potential. PMID:20838577